Increased primary carnitine deficiency detection through second-tier newborn genetic screening

Abstract

Background Newborn screening for (NBS) for primary carnitine deficiency (PCD) is widely implemented worldwide, however, with poor sensitivity. This study aimed to evaluate the feasibility of improving the screening using second-tier genetic assay. Methods An Agena iPLEX assay was developed to identify 17 SLC22A5 mutations in the Chinese populations, then this assay was applied in NBS for second-tier mutation screening of newborns with low free carnitine (C0) levels. Results The Agena iPLEX assay was successfully established and performed in second-tier genetic screening. A total of316 (0.15%) residual NBS-positive specimens were subjected to second-tier genetic screening. Twenty screen-positive newborns harbored biallelic mutations in SLC22A5 gene, 99 screen-carriers with one mutation, and 197 screen-negative newborns with no mutation identified. Among 99 carriers, four newborns were found with a second disease-causing SLC22A5mutation by further genetic analysis. Four newborns were found with persistently low C0 levels among 197 screen-negatives, further genetic analysis revealed that one newborn with two novel SLC22A5 pathogenic variants. In total, 25 newborns were diagnosed with PCD, for a positive predictive value of 7.91% (25/316). The incidence of PCD in Quanzhou was estimated at 1:8191. Conclusion Data from this study revealed that 24% (6/25) of PCD cases would have been missed by conventional NBS. The high throughput iPLEX assay is a powerful tool for PCD genotyping. The incorporation of second-tier genetic screening into NBS program could increase PCD detection, however, further studies are needed to optimize the workflow of new screening algorithm.

Biochemical And Genetic Characteristics of Patients With Primary Carnitine Deficiency Identified Through Newborn Screening

“…NBS results showed that all patients with low C0 levels, while some patients may have normal C0 levels during recall review. These patients would have been excluded from conventional NBS, but could be identi ed by incorporating second-tier genetic screening [19]. However, it is noteworthy that PCD patients are also easily missed during NBS because the C0 levels at birth could be affected by the maternal concentration and therefore cannot re ect the true levels of the newborns themselves.…”

“…Newborns who tested positive on the second screen were referred for further genetic analysis. Between January 2017 and December 2018, a second-tier newborn genetic screening targeting 17 SLC22A5 variants was added to our NBS program [19]. Targeted next-generation sequencing (NGS) was performed as previously described [27].…”

“…The mean C0 level at NBS in this cohort was 4.16 ± 1.78 µmol/L (range 1.63-8. 2021 [19] C0: free carnitine detected at newborn screening, C0-F1: C0 retested at recall stage, cutoff value: 8. 2021 [19] C0: free carnitine detected at newborn screening, C0-F1: C0 retested at recall stage, cutoff value: 8.…”

“…2021 [19] C0: free carnitine detected at newborn screening, C0-F1: C0 retested at recall stage, cutoff value: 8. 2021 [19] C0: free carnitine detected at newborn screening, C0-F1: C0 retested at recall stage, cutoff value: 8. 2019 [24] 41 Female 6.83 4.59 c.760C>T (p.R254*) c.1400C>G(p.S467C) Lin et al 2019 [24] 42 Female 6.22 11.14 c.760C>T (p.R254*) c.1400C>G(p.S467C) Lin et al 2019 [24] 43 Female 6.16 4.02 c.695C>T(p.T232M) c.1400C>G(p.S467C) Lin et al 2019 [24] 44 Male 6.77 4.5 c.760C>T (p.R254*) c.1400C>G(p.S467C) Lin et al 2019 [24] 45 Female 4.91 6.66 c.250T>A(p.Y84N) c.1400C>G(p.S467C) Lin et al 2019 [24] 46 Male 3.24 4.05 c.51C>G (p.F17L) c.1196G>A(p.R399Q) Lin et al 2019 [24] 47 Male 4.96 5 c.51C>G (p.F17L) c.1195C>T(p.R399W) Lin et al 2019 [24] 48 Female 3.15 4.09 c.760C>T (p.R254*) c.1400C>G(p.S467C) Lin et al 2019 [24] 49 Female 4.12 1.29 c.760C>T (p.R254*) c.760C>T (p.R254*) Lin et al 2019 [24] C0: free carnitine detected at newborn screening, C0-F1: C0 retested at recall stage, cutoff value: 8.5-50 µmol/L.…”