Expanded Newborn Screening for Detection of Vitamin B₁₂ Deficiency FREE

To the Editor: When undiagnosed, infant vitamin B₁₂ deficiency can result in anemia, failure to thrive, developmental regression, and neurological deficits and is most commonly caused by maternal vitamin B₁₂ deficiency.¹ Biochemically, vitamin B₁₂ deficiency leads to an accumulation of total homocysteine (tHcy), methylmalonic acid (MMA), and propionylcarnitine (C₃). Although vitamin B₁₂ deficiency is not a primary target of newborn screening (NBS) programs, measurement of C₃ may incidentally identify vitamin B₁₂–deficient newborns. However, first-tier screening is limited because C₃ levels may not be sufficiently high during the first days of life when most NBS samples are collected.² Levels of MMA and tHcy are more sensitive markers of vitamin B₁₂ status than serum B₁₂ levels in adults,³ have a strong inverse relationship with serum B₁₂ levels,⁴ and are usually elevated in infants who are vitamin B₁₂ deficient before hematologic or neurologic findings develop.² Through the public-private partnership between the Minnesota Department of Health, Mayo Clinic, and University of Minnesota, a lower first-tier cut-off level of C₃ was applied (>9.2 μmol/L to >5.25 μmol/L) and second-tier testing of MMA and tHcy levels in presumptive-positive NBS specimens was introduced in 2005.⁵

A population-based study of 363 649 infants born in Minnesota was conducted between January 1, 2005, and December 31, 2010. Newborn screening samples with elevated C₃ levels, a C₃ to acetylcarnitine (C₃/C₂) ratio of 0.1 or greater, or a C₃ to palmitoylcarnitine (C₃/C₁₆) ratio of 2 or greater had second-tier testing of tHcy (abnormal, >15 μmol/L) and MMA (abnormal, >5 μmol/L) on the same sample.⁵ (To convert tHcy to mg/dL, divide by 7.397.) All newborns with positive NBS results and some of their mothers underwent further laboratory testing. An additional newborn from another state whose first-tier NBS findings were normal presented at age 2 months with clinical complications of vitamin B₁₂ deficiency and required prolonged hospitalization. His original NBS sample was retrospectively analyzed to determine if the Minnesota 2-tier protocol would have led to identifying the infant as vitamin B₁₂ deficient. The University of Minnesota institutional review board approved review of medical records and waived informed consent.

Of 46 newborns with abnormal C₃ levels or related ratios identified in Minnesota, 11 had vitamin B₁₂ deficiency secondary to maternal vitamin B₁₂ deficiency (Table 1 and Table 2) for a detection rate of 3.02 per 100 000 live births; 22 findings were false positives; 1 declined follow-up testing; 1 had unconfirmed vitamin B₁₂ deficiency; 3 had cobalamin C defects; and 2 had methylmalonic acidemias. Five were homozygous and 1 a carrier for transcobalamin receptor defects. Newborn screening led to the identification of vitamin B₁₂ deficiency in 8 of 11 mothers whose status was previously unrecognized. All 11 newborns were asymptomatic when treatment was initiated (Table 2), and subsequently all biochemical markers normalized without clinical signs of vitamin B₁₂ deficiency. Retrospective analysis using the 2-tier protocol of the original NBS sample of the symptomatic infant screened by another state had positive results (Table 1, case 12).

A recent survey of 12 state NBS programs (including Minnesota) identified 32 cases of nutritional vitamin B₁₂ deficiency, for a rate of 0.88 per 100 000 newborns.⁶ After implementing a lower C₃ first-tier cut-off level and second-tier testing of MMA and tHcy levels in NBS specimens, the detection rate in Minnesota of vitamin B₁₂–deficient infants was considerably higher (3.02/100 000 live births). None of the 11 infants would have been identified by using the previous C₃ cut-off value, potentially explaining the reason no cases of infant vitamin B₁₂ deficiency were identified by NBS before 2005 in Minnesota.

Based on the Minnesota experience, vitamin B₁₂ deficiency occurs more frequently than is currently recognized. Given the important role vitamin B₁₂ plays in DNA synthesis and central nervous system function, and the difficulty in diagnosing vitamin B₁₂ deficiency in infancy because of nonspecific clinical findings, early identification and initiation of treatment are crucial to avoid potentially devastating and irreversible neurologic damage.