Evaluation of Single Nucleotide Variants in Intron 1 of the ABO Gene As Diagnostic Markers for the A Blood Group
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Introduction: The molecular diagnosis of the A blood group is based on the exclusion of gene variants causing blood groups A, B, or O. A specific genetic marker for the A blood group is still missing. Recently, long-read ABO sequencing revealed four sequence variations in intron 1 as promising markers for the * allele. Here, we evaluated the diagnostic values of the 4 variants in blood donors with regular and weak A phenotypes and genotypes.
Methods: ABO phenotype data (A, B, AB, or O) were taken from the blood donor files. The genotypes (low resolution) were known from a previous study and included the variants c.261delG, c.802G>A, c.803G>C, and c.1061delC. variant alleles (****, *, and *) were identified in weak A donors by sequencing the exons before. For genotyping of the intron 1 variants rs532436, rs1554760445, rs507666, and rs2519093, we applied TaqMan assays with endpoint fluorescence detection according to a standard protocol. Genotypes of the variants were compared with the ABO phenotype and genotype. Evaluation of diagnostic performance included sensitivity, specificity, positive (PPV), and negative predictive value (NPV).
Results: In 1,330 blood donors with regular ABO phenotypes and genotypes, the intron 1 variants were significantly associated with the proposed A blood group. In 15 donors, we found discrepancies to the genotype of at least one of the 4 variants. For the diagnosis of the ABO*A1 allele, the variants showed 98.79-99.48% sensitivity, 99.66-99.81% specificity, 98.80-99.31% PPV, and 99.66-99.86% NPV. Regarding the A phenotype, the diagnostic values were 99.02-99.41% sensitivity, 99.63-99.76% specificity, 99.41-99.61% PPV, and 99.39-99.63% NPV. The * marker allele of all intron 1 variants was also associated with the *, *, and * variants. Samples with *, *, and * variants lacked this association.
Conclusion: The intron 1 variants revealed significant association with the * allele and the A phenotype. However, the intron 1 genotype does not exclude variant alleles causing weak A phenotypes. With the introduction of reliable tag, single nucleotide variants for the A, A, B, and O blood groups and the genotyping instead of phenotyping of the ABO blood group are getting more feasible on a routine basis.