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DNA carrier testing and newborn screening for maple syrup urine disease in Old Order Mennonite communities
Stephanie M Carleton1, Dawn S Peck, Julie Grasela
1Department of Biochemistry, University of Missouri-Columbia, Columbia, Missouri 65211, USA.
Abstract:
Maple syrup urine disease (MSUD) is an inherited metabolic disorder caused by mutations in the branched chain alpha-keto acid dehydrogenase complex. Worldwide incidence of MSUD is 1:225,000 live births. However, within Old Order Mennonite communities, the incidence is 1:150 live births and results from a common tyrosine to asparagine substitution (Y438N) in the E1alpha subunit of branched chain alpha-keto acid dehydrogenase. We developed a new DNA diagnostic assay utilizing TaqMan technology and compared its efficacy, sensitivity, and duration with an existing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. Carrier testing was performed by both TaqMan technology and PCR-RFLP on DNA isolated from buccal swabs of 160 individuals as well as from buccal swabs and blood spots of nine at-risk newborns; assay time, sensitivity, and reliability were also evaluated. The TaqMan assay, like the PCR-RFLP assay, accurately determined Y438N E1alpha allele status. However, the TaqMan assay appeared (1) more sensitive than the PCR-RFLP assay, requiring 10-fold less DNA (10 ng) to reliably determine genotype status and (2) faster, reducing the assay time required for diagnosis from approximately 12 to 5 h. TaqMan technology allowed more rapid DNA diagnoses of MSUD in the neonate, thereby reducing the likelihood of neurological impairment while enhancing health and prognosis for affected infants.
Insights
Maple syrup urine disease (MSUD) is a rare inherited disorder. A new TaqMan DNA diagnostic assay offers faster and more sensitive detection of MSUD, improving infant health outcomes.
Area of Science:
- Genetics
- Biochemistry
- Medical Diagnostics
Background:
- Maple syrup urine disease (MSUD) is an inherited metabolic disorder affecting 1 in 225,000 live births globally.
- A specific mutation (Y438N) in the E1alpha subunit of branched-chain alpha-keto acid dehydrogenase is prevalent in Old Order Mennonite communities, increasing incidence to 1 in 150 live births.
Purpose of the Study:
- To develop and evaluate a novel DNA diagnostic assay using TaqMan technology for Maple syrup urine disease (MSUD).
- To compare the efficacy, sensitivity, and speed of the new TaqMan assay against the existing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for MSUD diagnosis.
Main Methods:
- Carrier testing and genotype determination were performed on DNA from buccal swabs of 160 individuals and from newborns using both TaqMan and PCR-RFLP assays.
- Key performance metrics including assay time, sensitivity, and reliability were evaluated for both diagnostic methods.
Main Results:
- The TaqMan assay demonstrated high accuracy in determining the Y438N E1alpha allele status, comparable to the PCR-RFLP assay.
- The TaqMan assay required 10-fold less DNA (10 ng) and reduced assay time from 12 hours to 5 hours, indicating superior sensitivity and speed.
- The new assay enables more rapid DNA diagnoses of MSUD in neonates.
Conclusions:
- TaqMan technology provides a more sensitive and significantly faster diagnostic method for MSUD compared to PCR-RFLP.
- Rapid diagnosis of MSUD using TaqMan technology can reduce the risk of neurological impairment and improve health prognosis for affected infants.
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