Investigation of discrepancies obtained during 15 years of non‐invasive fetal RHD genotyping in apparent serologic RhD‐negative pregnant women

Abstract

Objectives In some European countries, non‐invasive fetal RHD genotyping is the first step of anti‐D allo‐immunized pregnant women management but presence of RHD variant alleles may interfere with the results accuracy. We developed an algorithm allowing solving discordant results (due to the presence of RHD variant) in fetal RHD genotyping assay. Method This study gathered the results of fetal RHD genotyping performed between 2006 and 2020 in the Medicine Laboratory of CHR Liège. Exons 4, 5 and 10 of the fetal RHD were profiled in maternal plasma using real time polymerase chain reaction (PCR). When the results were discrepant, maternal RHD variant was further explored by sequence‐specific primer PCR on maternal buffy coat. Results A total of 11,630 pregnant women (mainly of both Caucasian and African origins) were tested during the study period and RHD variant alleles were detected in 247 women. The most frequent variant was RHD*08N.01 found in 66 women mainly of Black African origin. We identified 45 women with weak RHD variant type 1, 2 or 3. Conclusion Women with weak RHD variant type 1, 2 or 3 can safely be considered as RhD positive in terms of RhIg prophylaxis and/or transfusion of blood components. Therefore, identification of RHD allele variants in women with discordant fetal RHD genotyping results contributes to save RhIg prophylaxis and RhD negative blood components.

Noninvasive screening of fetal RHD genotype in Chinese pregnant women with serologic RhD‐negative phenotype

“…Another possible reason is associated with a higher frequency of RHD variant alleles among Chinese negative pregnant women. In Caucasian, about 95% of individuals with the RhD‐negative phenotype resulting from homozygous RHD deletion ( RHD*01N.01 / RHD*01N.01 ), and the fetal RHD genotype can be predicted by detecting the presence or absence of the RHD gene in maternal cell‐free DNA 10,11 . However, more than 30% of the Chinese RhD‐negative phenotype is associated with variant RHD alleles, such as c.1227G>A ( RHD*01EL.01 ) and RHD‐CE‐D hybrid ( RHD * 01N.03 ) 12,13 .…”

“…In Caucasian, about 95% of individuals with the RhD-negative phenotype resulting from homozygous RHD deletion (RHD*01N.01/RHD*01N.01), and the fetal RHD genotype can be predicted by detecting the presence or absence of the RHD gene in maternal cell-free DNA. 10,11 However, more than 30% of the Chinese RhD-negative phenotype is associated with variant RHD alleles, such as c.1227G>A (RHD*01EL.01) and RHD-CE-D hybrid (RHD*01N.03). 12,13 Relatively high frequency of variant RHD alleles in pregnant women would limit the accuracy of fetal RHD status prediction due to the fact that fetal cell-free DNA only accounts for 10%-15% of total cell-free DNA.…”

Misclassification of RhD variants among pregnant women: a systematic review

“…These terminologies surrounding RhD variants lead to confusion in RhD result interpretation. In prenatal tests, pregnant women grouped as RhD- are considered eligible to receive RhD immunoglobulin (RhIG) as a preventive measure for HDFN [ 12 , 13 ]. However, some women who are RhD are mistakenly grouped as RhD+ or weak D and are not eligible for receiving RhIG, which puts them at risk of RhD alloimmunization and their fetuses at risk of HDFN.…”