The RHD gene is highly polymorphic and the existence of a large number of alleles results in RhD variant phenotypes. RHD genotyping has been used to distinguish normal D antigen from D variants due to limitations of serologic methods. The purpose of this study was to determine the phenotypic frequency of RhD and RhCE antigens and to investigate the RHD alleles present in samples with the weak D or D- phenotypes from Brazilian blood donors. A total of 2007 donors were phenotyped for D, C, c, E and e antigens. Samples phenotyped as D- were genotyped by polymerase chain reaction-sequence specific primers, and exon 10 and intron 4 of the RHD gene were analysed. D- samples containing the RHD gene or samples considered weak D were further characterised using genotyping platform or nucleotide sequencing. Using serologic methods we found that 87.3% of the donors were D+, 11.9% D- and 0.8% weak D. The frequency of RHD gene in D- individuals was 9.2%. Five RHD alleles from phenotypically D- donors were characterised in six molecular backgrounds: RHDΨ, RHD-CE-D(s), RHD-CE-(2-9)-D, RHD/RHDΨ, RHDΨ/RHD-CE-D(s) and RHD-CE(2)-D. The most common weak D antigens types found were 1, 3, 4.0/4.1 and 4.2, whereas the most prevalent weak D type was 4.2 (or DAR). The RHD genotyping proved to be a necessary tool to characterise RHD alleles in donors phenotyped as D- or weak D to increase the transfusion safety in highly racial mixed population.
Background and Objectives
The high homology and the inverted orientation of RHD and RHCE may give rise to non‐functional and aberrant RH alleles. RH genotyping is used to screen RH matched donors to African descent patients. This study aimed to define a strategy for testing RHD and RHCE variants in blood donors to provide compatible units for transfusion of patients with haematological diseases.
Materials and Methods
Samples from 132 patients [101 Sickle cell disease (SCD), 14 myelodysplastic syndrome (MDS), 17 acute myelogenous leukaemia (AML)] and 198 Brazilian donors were studied. Major blood group alleles, RHD, RHCE alleles and RHD zygosity were determined by the blood‐MLPA assay. Sequencing was performed to determine RHD and RHCE variant subtypes. A match was an RH genotype that did not encode Rh antigens absent in the patient, along with matching for ABO, MNS, KEL, FY, JK and DI antigens.
Results
Overall, 7·6% of blood donors and 17.4% of patients presented RH genotypes that predict expression of partial Rh antigens or lack of high prevalence Rh antigens. From 23 patients with clinically relevant RH genotypes, 15 had available matched donors.
Conclusion
We report the presence of clinically relevant RH genotypes in SCD and in non‐SCD patients. In our admixed population, many patients carry variant RHCE alleles in heterozygosity with normal RHCE alleles. Thus, our results suggest that donors could be selected based on the normal RH allele.
The results demonstrate the importance of the clinical analyst's knowledge about the behavior of the CBC parameters over time under different storage conditions, and mainly the imprecision of the hematological equipment used, for the suitable interpretation of the results.
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