Detection of interferon alpha and beta receptor subunit 1 (IFNAR1) loss-of-function Glu386∗ variant by tri-allelic genotyping.

Mutations of the human interferon alpha and beta receptor subunit 1 (IFNAR1) gene are associated with severe viral infections. Individuals homozygous for the Glu386∗ variant have impaired type I interferon signalling and can suffer severe illness when exposed to certain viruses and live attenuated virus vaccines. Glu386∗ heterozygotes are clinically unaffected, but can pass the variant allele to their descendants. We aimed to develop an assay that can identify IFNAR1 Glu386∗ homozygotes and heterozygotes to support urgent clinical diagnosis, and that can use dried blood spots (DBS) sent at ambient temperature to overcome geographical logistical challenges in the South Pacific region. The tri-allelic genotyping assay interrogates a single nucleotide polymorphism (rs201609461) located in IFNAR1. The reference allele G encodes for wild-type IFNAR1. Minor alleles A (c.1156G>A) and T (c.1156G>T) encode for Glu386Lys and a truncated IFNAR1 protein (p.Glu386∗), respectively. Synthetic oligonucleotides were mixed in equal molar ratio to create six different genotypes which were randomly assigned to 960 genotyping reactions by R software. Three different fluorescence probes were designed to discriminate the three alleles (G, T and A) within a pair of flanking primers in a single genotyping reaction. The assay discriminated all three alleles using DBS from Guthrie cards randomly spiked with synthetic oligonucleotides. We correctly identified all the different genotypes in 960 reactions in these blinded experiments. These findings validate the genotyping assay for rapidly identifying the IFNAR1 Glu386∗ variant from DBS.

Introducing Newborn Screening for Severe Combined Immunodeficiency-The New Zealand Experience.

Screening for severe combined immunodeficiency (SCID) was added to the New Zealand national newborn screening programme in December 2017. Documentation pertaining to the application to add SCID to the panel and screening results over the first three years were reviewed. Screening evaluation metrics were shown to differ according to site of collection (babies in a neonatal intensive care unit vs. the community), definition of a positive test (out-of-range result vs. result leading to a further action on baby), and screening target/case definition (primary SCID vs. non-SCID T-cell lymphopenia). Our experience demonstrates both the value of close clinical involvement during the implementation phase of SCID screening and that the use of standard definitions will facilitate international comparison.