Dataset Information

RHD variants in Polish blood donors routinely typed as D-.

ABSTRACT:

Background

Blood donors exhibiting a weak D or DEL phenotypical expression may be mistyped D- by standard serology hence permitting incompatible transfusion to D- recipients. Molecular methods may overcome these technical limits. Our aim was to estimate the frequency of RHD alleles among the apparently D- Polish donor population and to characterize its molecular background.

Study design and methods

Plasma pools collected from 31,200 consecutive Polish donors typed as D- were tested by real-time polymerase chain reaction (PCR) for the presence of RHD-specific markers located in Intron 4 and Exons 7 and 10. RHD+ individuals were characterized by PCR or cDNA sequencing and serology.

Results

Plasma cross-pool strategy revealed 63 RHD+ donors harboring RHD*01N.03 (n = 17), RHD*15 (n = 12), RHD*11 (n = 7), RHD*DEL8 (n = 3), RHD*01W.2 (n = 3), RHD-CE(10) (n = 3), RHD*01W.3, RHD*01W.9, RHD*01N.05, RHD*01N.07, RHD*01N.23, and RHD(IVS1-29G>C) and two novel alleles, RHD*(767C>G) (n = 3) and RHD*(1029C>A). Among 47 cases available for serology, 27 were shown to express the D antigen

Conclusion

1) Plasma cross-pool strategy is a reliable and cost-effective tool for RHD screening. 2) Only 0.2% of D- Polish donors carry some fragments of the RHD gene; all of them were C or E+. 3) Almost 60% of the detected RHD alleles may be potentially immunogenic when transfused to a D- recipient.

SUBMITTER: Orzinska A

PROVIDER: S-EPMC5497842 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Publications

RHD variants in Polish blood donors routinely typed as D-.

Orzińska Agnieszka A Guz Katarzyna K Polin Helene H Pelc-Kłopotowska Monika M Bednarz Justyna J Gieleżyńska Agata A Sliwa Beata B Kowalewska Małgorzata M Pawłowska Elżbieta E Włodarczyk Bogusława B Malaga Alicja Żmudzin Małgorzata M Krzemienowska Magdalena M Srivastava Kshitij K Michalewska Bogumiła B Gabriel Christian C Flegel Willy A WA Brojer Ewa E

Transfusion 20130501 11 Suppl 2

<h4>Background</h4>Blood donors exhibiting a weak D or DEL phenotypical expression may be mistyped D- by standard serology hence permitting incompatible transfusion to D- recipients. Molecular methods may overcome these technical limits. Our aim was to estimate the frequency of RHD alleles among the apparently D- Polish donor population and to characterize its molecular background.<h4>Study design and methods</h4>Plasma pools collected from 31,200 consecutive Polish donors typed as D- were teste ...[more]

PMID: 23634715

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Project description:In the 1980s, Poland was a medium-endemic country, with one of the highest incidences of hepatitis B in Europe (45/105 inhabitants). Pursuant to the WHO guidelines, obligatory vaccination was introduced in 1994-1996 (as a part of hepatitis B prophylaxis for newborns), and in 2000-2011, all 14-year-olds were vaccinated. To prevent transfusion-transmitted HBV infection (TT-HBV), since the 1970s, each donation has been tested for HBsAg and, since 2005, additionally for the presence of HBV DNA. Based on the data from the Blood Transfusion Centers, changes in HBV detection in Polish blood donors were analyzed, starting from the introduction of mandatory NAT screening until 2019. During the period under analysis, a total of 11,625 HBV-infected donors were identified: 97.95% were seropositive (confirmed HBsAg) and 2.05% were seronegative (NAT yields). The detection frequency for both categories of infections was significantly (p = 0.05) higher for men than for women (Residual Risk RR = 1.4 and RR = 2.63, respectively). Seropositive infections were detected more frequently (p < 0.05) in first-time donors than in repeat donors (RR = 360), while no significant differences were observed in the category of seronegative infections. A downward trend in HBsAg detection was observed in both first-time and repeat donors (Spearman's coefficient R = -0.98 and R = -0.90, respectively). The frequency of HBsAg in first-time donors decreased 5-fold, and, in repeat donors, 30-fold. In both subpopulations, the largest decrease occurred in the age group ≤ 20 years (i.e., donors born between 1985 and 2001). The incidence of window period (WP) infections in the repeat donor group demonstrated a downward trend (R = -0.54, p < 0.05), and in the first-time donor group, no significant trend was recorded. For occult hepatitis B infection (OBI), no significant trend was observed in either donor subpopulation. WP infections were detected significantly more often in donors aged 21-50 years than in donors ≤20 years, most often in the 41-50 age group. The frequency of OBI increased with donor age and was the highest in the 51-60 age group. A spectacular decrease in the frequency of HBsAg(+) infections was observed in current study, indicating the effectiveness of the hepatitis prevention strategy applied in Poland. We expect that the improvement in the epidemiological situation among blood donors causes a reduction in the risk of TT-HBV. Confirmation of this hypothesis by the analysis of residual risk should be a subject of further studies.

Project description:BackgroundHybrid genes are responsible for the formation of Rh variants and are common in patients with sickle cell disease (SCD). However, it is not usually possible to detect them by conventional molecular protocols. In the present study, hybrid genes were investigated using the Quantitative Multiplex Polymerase chain reaction of Short Fluorescent Fragments (QMPSF), a molecular protocol that quantifies the copy number of RHD and RHCE exons. In addition, we explored additional relevant information obtained with QMPSF, such as recognition of variant RHCE and RHD zygosity.Materials and methodsThree groups of subjects were selected for the study: patients with SCD, self-declared African descent donors (SDA), and D-negative donors. RHD and RHCE hybrids genes were investigated by the QMPSF method. Real-time multiplex polymerase chain reaction (PCR) assay was used to confirm the copy number of the RHD in two samples. Cloning was performed to investigate the allele. Relative RhD antigen density was investigated by flow cytometry, and RhCE phenotyping was performed with both tube and gel methods.ResultsIn the 507 samples analysed, hybrid allele frequencies were found in 20.08% of patients with SCD, in 18.22% of individuals in the SDA group, and 3.67% of D-negative donors. The SCD and SDA groups had a higher frequency of hybrid alleles, most commonly involving exon 8, with which we found an association with c.733C>G, a common polymorphism observed in individuals of African descent. Of note, two patients with SCD were shown to carry three gene copies, as confirmed by quantitative PCR; no increase in D expression was observed in these patients. In addition, the QMPSF guided the investigation of 144 RHCE variants and RHD zygosity, and two novel alleles were identified.DiscussionThe QMPSF was shown to identify hybrid alleles involved in altered Rh phenotypes in Brazilian donors and patients with SCD. The association of the hybrid RHCE-D(8)-CE allele with c.733C>G suggests this hybrid allele may be used as a marker to detect the most frequent variants found in patients with SCD.

Dataset Information

RHD variants in Polish blood donors routinely typed as D-.

Background

Study design and methods

Results

Conclusion

Publications

RHD variants in Polish blood donors routinely typed as D-.

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