Diagnostic Challenges Using a 2-Tier Strategy for Methylmalonic Acidurias: Data from 1.2 Million Dried Blood Spots

Abstract

<b><i>Background:</i></b> The detection of methylmalonic acid (MMA) by second-tier analysis has been shown to reduce the number of false positives in newborn screening (NBS) for genetically determined methylmalonic acidurias (MMAuria). In addition to genetic conditions, MMA is an indicator of vitamin B12 status, thus applicable to detect maternal vitamin B12 deficiency in the newborns screened. <b><i>Methods:</i></b> Biochemical and clinical follow-up data of a 7.5-year pilot study with 1.2 million newborns screened were reviewed. <b><i>Results:</i></b> Among 1,195,850 NBS samples, 3,595 (0.3%) fulfilled criteria for second-tier analysis of MMA. In 37 (0.003%; 1/32,000) samples, elevated concentrations of MMA were detected, resulting in diagnostic workup at a metabolic center in 21 newborns. In 6 infants (1/199,000), genetic conditions were established, 1 infant with cobalamin C deficiency (CblC) showed only a moderate elevation of MMA. The remaining 15 newborns (1/79,000) displayed significantly lower concentrations of MMA and were evaluated for maternal vitamin B12 deficiency. In 9 mothers, vitamin B12 deficiency was verified, and 6 showed no indication for vitamin B12 deficiency. Treatment with vitamin B12 normalized biochemical parameters in all 15 infants. <b><i>Conclusions:</i></b> Applying a 2-tier strategy measuring MMA in NBS identified genetic conditions of MMAuria. It was possible to separate severe, early-onset phenotypes from maternal vitamin B12 deficiency. However, the detection of CblC deficiency with mildly elevated MMA interferes with impaired vitamin B12 status of unknown relevance and thus burdens possibly healthy newborns. Regarding maternal vitamin B12 deficiency, testing and supplementing mothers-to-be is preferable. This might decrease straining follow-up of newborns and improve quality and overall perception of NBS.

Implementation of second-tier tests in newborn screening for the detection of vitamin B12 related acquired and genetic disorders: results on 258,637 newborns

“…In agreement with Gramer et al [24] Hcys is the best second-tier biomarker for the detection of acquired vitamin B 12 deficiency (Table 2B) and could explain why other programs [20,22,25] have found a low number of vitamin B 12 deficiencies since they do not use Hcys as second-tier test. In agreement with Hawthorne et al reflections [37], the measurement of Hcys as secondtier test is an economical way to increase the number of vitamin B 12 deficient infants identified, but also of some treatable genetic disorders.…”

“…Recently, an incidence of 1:3,000 newborns in the Estonian screening program has been reported [23], while another recent German study of Munich NBS program revealed a much lower incidence of vitamin B 12 deficiency [25]. Therefore, the incidence of this deficiency differs considerably among NBS programs and it might reflect different strategies used in each program, or even dietetic cultural factors dependent on the nationalities of origin, or other demographic factors.…”

“…Therefore, the incidence of this deficiency differs considerably among NBS programs and it might reflect different strategies used in each program, or even dietetic cultural factors dependent on the nationalities of origin, or other demographic factors. On the other hand, the Italian, Estonian and Municher programs [22,23,25] are mainly based on the use of C3 and MMA, while our program and the German pilot screening of Heidelberg [24] are based on the detection of metabolites of both cobalamin dependent pathways. In addition, the cut-offs used play a relevant role, as exemplified by our two strategies.…”

Vitamin B12 deficiency in newborns and infants—Causes, early detection, diagnostics and presentation of a primary oral treatment scheme

“…Hierzu verfassten wir einen Kommentar im Leserforum zur Möglichkeit der Früherkennung des Vitamin-B12-Mangels im Neugeborenenscreening im Rahmen von aktuellen Pilotprojekten [10]. Mittlerweile wurden im Rahmen solcher Projekte bereits zahlreiche Kinder mit maternal bedingtem Vitamin-B12-Mangel identifiziert und früh behandelt [8,22,26]. Mit einer Inzidenz von bis zu einem betroffenen Kind unter 5300 untersuchten Neugeborenen war im Pilotprojekt "Neugeborenenscreening 2020" am Screeningzentrum Heidelberg der maternal bedingte Vitamin-B12-Mangel häufiger als jede andere der metabolischen Zielkrankheiten des aktuellen Regelscreenings in Deutschland [4,8,12].…”

Newborn Screening for Methylmalonic Acidemia/propionic Acidemia: Systematic Evaluation of a Two-pronged Approach Based on MS/MS and UPLC-MS/MS

“…C3associated disorder screening is fraught with false-positive cases that require second-tier con rmation using UPLC-MS/MS, and DNA sequencing is necessary to reach a nal diagnosis and to identify which pathogenic genes are responsible and the severity of the speci c variant. On the basis of 509,313 newborns born in Xuzhou, China, over a 5-year period between November 2015 and December 2020 C3, C3/C2 and methionine (Met) data measured by rst-tier MS/MS technologies, we developed a novel strategy using MMA, methylcitrate (MCA) and homocysteine (HCY) values as second-tier test parameters for differential screening in initially positive samples 14 . In addition to the primary MS/MS analytes currently used in NBS for MMAs and PA, our statistical approach utilizes information from the entire MS/MS metabolic pro le measured at birth.…”