Sickle cell disease patients in two London trusts: Genotyping including RH variants.

BACKGROUND: All SCD patients need extended RBC antigen typing (by serology or genotyping) for provision of extended RH, K matched blood and to guide RBC selection in those with complex transfusion requirements. Genotyping can also identify RH variants which can cause sensitisation even when extended RH phenotypically matched blood is provided and alloantibodies associated with RH variants can cause HTRs. OBJECTIVES: To review the use of RBC genotyping in SCD patients at two London trusts (ICHNT, LNWH) with a focus on RH variants. METHODS: Retrospective review with data collected from clinical notes, local and national pathology reporting systems. RESULTS: A 311/482 (64%) ICHNT patients and 181/346 (52%) LNWH patients had extended genotyping. Of genotyped patients, 68 (22%) ICHNT and 31 (17%) LNWH patients had RH variants. Eight ICHNT patients had RH variants and corresponding antibodies associated with RH variants; 4/8 received multiple transfusions with antigen positive RBCs but had no evidence of haemolysis. One LNWH patient had a RH variant with corresponding alloantibody but could not be investigated further for possible HTR. CONCLUSIONS: Most patients (59%) had genotyping and a significant number had RH variants (99, 20%). A small proportion (9, 9%) had antibodies associated with RH variants, but with no evidence of clinically significant HTRs despite transfusions in four of them with antigen positive RBCs. All SCD patients should have RBC genotyping including RH variants (preferentially over extended phenotyping) to guide better selection of RBC units. However, where antigen negative blood cannot be provided, the risk of alloimmunisation is not inevitable and subsequent HTRs from antibodies associated with RH variants might not always occur.

Characterization of GYP*Mur and novel GYP*Bun-like hybrids in Thai blood donors reveals a qualitatively altered s antigen.

BACKGROUND AND OBJECTIVES: The Mi(a+) GP(B-A-B) hybrid phenotypes occur with a prevalence of 2%-23% across Southeast Asia. While the s antigen is alleged to be altered, no evidence for specific variants is known. Screening using a monoclonal IgM anti-s mistyped six S-s+ RBC units as S-s-. Further, alloanti-s was identified in an S+s+ patient. Our objective was to investigate the s antigen further. MATERIALS AND METHODS: DNA from 63 Thai blood donor samples PCR-positive for a GYP(B-A-B) hybrid was sequenced with primers spanning GYPB exons 3-4. Flow cytometry was used for semiquantitative analysis of s expression and correlated with the glycophorin genotype. RESULTS: DNA sequencing showed that GYP*Mur was carried by 56/63 samples (88·9%) of which 5/56 lacked normal GYPB: three of these were GYP*Mur homozygotes, one was a compound heterozygote carrying GYP*Mur and a GYP*Bun-like allele (designated GYP*Thai), and the fifth sample carried GYP*Mur and another GYP*Bun-like allele. Seven samples (7/63) were GYP*Thai heterozygotes. IgM monoclonal anti-s (P3BER) did not react with the s antigen carried by GP.Mur or GP.Bun, whereas two IgG anti-s showed enhanced reactivity. CONCLUSIONS: We confirmed that GYP*Mur is the most frequent variant in Thai blood donors and also identified GYP*Thai with a frequency of 1·1%. We showed that s antigen on Mi(a+) GP(B-A-B) hybrids is qualitatively altered and should be considered when selecting reagents for phenotyping where such hybrids are prevalent, endemically and in blood centres worldwide.

Clinical significance of antibodies to antigens in the ABO, MNS, P1PK, Rh, Lutheran, Kell, Lewis, Duffy, Kidd, Diego, Yt, and Xg blood group systems.

CONCLUSIONS: This report is part of a series reporting the proceedings from the International Society of Blood Transfusion (ISBT) Working Party on Immunohaematology Workshop on the Clinical Significance of Red Blood Cell Alloantibodies. The aim of the workshop was to review information regarding the clinical significance of alloantibodies to red blood cell antigens recognized by the ISBT. The first 12 systems will be covered in this report. It is understandable that many of the most clinically important antibodies are directed toward antigens found in the blood group systems discovered earlier in history. The ABO system was the first to be discovered and remains the most clinically important regarding transfusion.

Lack of the nucleoside transporter ENT1 results in the Augustine-null blood type and ectopic mineralization.

The Augustine-negative alias At(a-) blood type, which seems to be restricted to people of African ancestry, was identified half a century ago but remains one of the last blood types with no known genetic basis. Here we report that a nonsynonymous single nucleotide polymorphism in SLC29A1 (rs45458701) is responsible for the At(a-) blood type. The resulting p.Glu391Lys variation in the last extracellular loop of the equilibrative nucleoside transporter 1 (ENT1; also called SLC29a1) is known not to alter its ability to transport nucleosides and nucleoside analog drugs. Furthermore, we identified 3 individuals of European ancestry who are homozygous for a null mutation in SLC29A1 (c.589+1G>C) and thus have the Augustine-null blood type. These individuals lacking ENT1 exhibit periarticular and ectopic mineralization, which confirms an important role for ENT1/SLC29A1 in human bone homeostasis as recently suggested by the skeletal phenotype of aging Slc29a1(-/-) mice. Our results establish Augustine as a new blood group system and place SLC29A1 as a new candidate gene for idiopathic disorders characterized with ectopic calcification/mineralization.

Is Next Generation Sequencing the future of blood group testing?

Blood group genotyping has many advantages over conventional phenotyping for both blood donors and patients, and a number of high-throughput methods have now been developed. However, these are limited by a requirement for existing knowledge of the relevant blood group gene polymorphisms, and rare or novel mutations will not be detected. These mutations could be successfully identified by DNA sequencing of the blood group genes, and such an approach has been made feasible by the introduction of Next Generation Sequencing (NGS) technology. NGS enables many genes from multiple samples to be sequenced in parallel, resulting in sequencing information that could be used to obtain accurate blood group phenotype predictions in both blood donors and patients.

Development and validation of a universal blood donor genotyping platform: a multinational prospective study.

Each year, blood transfusions save millions of lives. However, under current blood-matching practices, sensitization to non-self-antigens is an unavoidable adverse side effect of transfusion. We describe a universal donor typing platform that could be adopted by blood services worldwide to facilitate a universal extended blood-matching policy and reduce sensitization rates. This DNA-based test is capable of simultaneously typing most clinically relevant red blood cell (RBC), human platelet (HPA), and human leukocyte (HLA) antigens. Validation was performed, using samples from 7927 European, 27 South Asian, 21 East Asian, and 9 African blood donors enrolled in 2 national biobanks. We illustrated the usefulness of the platform by analyzing antibody data from patients sensitized with multiple RBC alloantibodies. Genotyping results demonstrated concordance of 99.91%, 99.97%, and 99.03% with RBC, HPA, and HLA clinically validated typing results in 89 371, 3016, and 9289 comparisons, respectively. Genotyping increased the total number of antigen typing results available from 110 980 to >1 200 000. Dense donor typing allowed identification of 2 to 6 times more compatible donors to serve 3146 patients with multiple RBC alloantibodies, providing at least 1 match for 176 individuals for whom previously no blood could be found among the same donors. This genotyping technology is already being used to type thousands of donors taking part in national genotyping studies. Extraction of dense antigen-typing data from these cohorts provides blood supply organizations with the opportunity to implement a policy of genomics-based precision matching of blood.

Resistance to malaria through structural variation of red blood cell invasion receptors.

The malaria parasite Plasmodium falciparum invades human red blood cells by a series of interactions between host and parasite surface proteins. By analyzing genome sequence data from human populations, including 1269 individuals from sub-Saharan Africa, we identify a diverse array of large copy-number variants affecting the host invasion receptor genes GYPA and GYPB We find that a nearby association with severe malaria is explained by a complex structural rearrangement involving the loss of GYPB and gain of two GYPB-A hybrid genes, which encode a serologically distinct blood group antigen known as Dantu. This variant reduces the risk of severe malaria by 40% and has recently increased in frequency in parts of Kenya, yet it appears to be absent from west Africa. These findings link structural variation of red blood cell invasion receptors with natural resistance to severe malaria.