Serological and molecular characterisation of the most prevalent weak D variants in Croatian population.

Introduction: Existence of hundreds of RHD gene variants contributes to variable D antigen expression and inconsistencies in reporting the RHD results. The aim of the study was to determine the serological and molecular characteristics of the most prevalent RHD alleles encoding serologically weak D variants.

Material And Methods: Blood donors (n = 145 924) were typed for D antigen using the direct serologic micromethod. Nonreactive samples were analysed in IAT method with the IgM/IgG anti-D monoclonal blend, and 0,2% (n = 263) confirmed weak D antigen expression. After genomic DNA extraction (Qiaqen, Germany), RHD genotyping was performed using in house reagents and PCR-SSP kits (Inno-Train, Germany).

Results: The prevalence of serologically weak D in blood donor population was 0.2% (n = 263). RHD genotyping confirmed weak D allele in 92.4% and partial D allele in 7.6%. The most common was weak D type 1 (49.7%) followed by weak D type 3 (24.7%) and type 2 (9.5%). Relatively high frequency was detected for weak D type 14 (4.6%) and type 64 (2.3%). In the category of partial D phenotypes, only DVI variant was found. Direct typing has shown great variability in the strength of reactions with different clones of anti-D reagents.

Conclusion: Weak D type 1 is the most common weak D variant in Croatian blood donor population. The frequency of D variants and distribution of Rh phenotypes in our study was in concordance with other studies. It has been shown that serological methods and the combination of clones used, cannot distinguish variant D types, which justifies the use of molecular methods.