Integrated serological and molecular investigation of ABO subgroups underlying atypical ABO typing in southeastern Brazil

Abstract Background and Objectives Numerous A and B subgroups can cause discrepancies between forward and reverse typing, weak reactivity or unexpected antibody patterns. This study aimed to identify the most prevalent ABO subgroups in a population of Southeast Brazil, characterize their serological profiles and evaluate their impact on ABO typing. Study Design and Methods Donor samples collected in São Paulo State, Brazil, were included in the study and classified into two groups according to predefined selection criteria. Group 1 comprised 27 samples with reduced or negative reactivity with anti‐A1 lectin, selected from 225 donors with routine Group A typing. Group 2 comprised 33 samples selected over a 2‐year period due to atypical ABO typing and subsequently subjected to complementary serological testing. All samples were molecularly characterized using polymerase chain reaction restriction fragment length polymorphism (PCR‐RFLP) and Sanger sequencing. Results In Group 1, the predominant allele was ABO*A2.01 , representing 10.2% of all type A donors. In Group 2, the most frequent variant allele was ABO*AW.09 (33%), followed by ABO*AW.31 (11%) and ABO*AEL.01 (11%). All these three subgroups were implicated in cases of ABO mistyping. B subgroups were identified in two samples and were associated with molecular changes in intron 1 (+5.8‐kb site). Conclusion Although ABO subgroups are uncommon, they occur in our population and directly affect blood typing. The results demonstrate the presence of clinically significant variants and highlight the need for vigilance in donor screening, including careful interpretation of typing results, the use of complementary serological testing and molecular analysis when necessary.