SID: a new carbohydrate blood group system based on a well-characterized but still mysterious antigen of great pathophysiologic interest.

The high-prevalence blood group antigen, Sda, had been puzzling blood bankers and transfusionists for at least a decade when it was reported in 1967. The characteristic mix of agglutinates and free red blood cells (RBCs), caused by anti-Sda, is seen with the RBCs from 90 percent of individuals of European descent. However, only 2-4 percent of individuals are truly Sd(a-) and may produce anti-Sda. The antibodies, generally considered insignificant, may cause hemolytic transfusion reactions with high-expressing Sd(a+) RBCs (e.g., the unusual Cad phenotype, which can also be polyagglutinable). The Sda glycan, GalNAcß1-4(NeuAcα2-3)Gal-R, is produced in the gastrointestinal and urinary systems, while its origin on RBCs is more controversial. According to current theory, Sda is likely to be passively adsorbed in low amounts, except in Cad individuals, where it has been found on erythroid proteins and at higher levels. The long-standing hypothesis that B4GALNT2 encodes the Sda synthase was confirmed in 2019, since homozygosity for a variant allele with rs7224888:C produces a non-functional enzyme associated with most cases of the Sd(a-) phenotype. Thereby, the SID blood group system was acknowledged as number 038 by the International Society of Blood Transfusion. Although the genetic background of Sd(a-) was settled, questions remain. The genetic background of the Cad phenotype has not yet been determined, and the source of the RBC-carried Sda is unknown. Furthermore, the interest of Sda stretches beyond transfusion medicine. Some tantalizing examples are lowered antigen levels in malignant tissue compared with normal tissue and interference with infectious agents like Escherichia coli, influenza virus, and malaria parasites.

Yes, MAM: how the cancer-related EMP3 protein became a regulator of erythropoiesis and the key protein underlying a new blood group system.

The MAM blood group system (International Society of Blood Transfusion [ISBT] 041) consists of one high-prevalence antigen to date, first detected in a 31-year-old woman during her third pregnancy. Epithelial membrane protein 3 (EMP3) was recently identified as the gene coding the MAM antigen. Six unique genetic variants have been described in EMP3 in 11 MAM- individuals. EMP3 is an 18-kDa glycoprotein with a large extracellular domain containing at least one N-glycosylation site. The normal function of EMP3 is still unclear, but ex vivo culture of erythropoietic progenitor cells from MAM- individuals shows an increased yield of reticulocytes, suggesting that EMP3 acts as a brake during normal erythropoiesis. EMP3 is abundant on different cell types, including many epithelial tissues and blood cells. Interestingly, EMP3 expression has been suggested as a prognostic marker for a number of cancer types, both for good and poor prognoses. EMP3 may act as a tumor suppressor or an oncogene in different cancer contexts. The protein appears to interact with other cell surface receptors and affects the downstream signaling and function of these proteins. MAM- red blood cells express low levels of CD44 and, consequently, the antigens of the Indian blood group system are only weakly expressed. Clinically, the MAM blood group antigen is important with regard to blood transfusion and pregnancy. Anti-MAM can cause severe hemolytic disease of the fetus and newborn in some pregnancies but have little to no effect in other pregnancies. Cases are typically not detected until problems occur during pregnancy, making the availability of compatible blood a challenge.

Multiple miscarriages in two sisters of Thai origin with the rare Pk phenotype caused by a novel nonsense mutation at the B3GALNT1 locus.

OBJECTIVES: To determine the genetic background underlying the Pk phenotype in two Thai sisters suffering from multiple spontaneous abortions. BACKGROUND: The P antigen is carried by globoside, an abundant glycosphingolipid in the red blood cell (RBC) membrane. Inactivating mutations in the 3-ß-N-acetylgalactosaminyltransferase gene (B3GALNT1) give rise to the rare Pk phenotype, which lack the P and PX2 antigens. Consequently, naturally occurring anti-P may cause recurrent miscarriages following the cytotoxic attack of the globoside-rich fetal portion of the placenta. METHODS/MATERIALS: P/P1/PX2/Pk antigens on RBCs and their corresponding antibodies were detected by haemagglutination and flow cytometry. The B3GALNT1 coding region was sequenced, and an allele-specific polymerase chain reaction (PCR) was developed. RESULTS: The two sisters had suffered 8 and 11 miscarriages, most of which occurred in the first trimester. Anti-P and anti-PX2 were identified in their plasmas, and the RBCs typed as P-PX2-Pk +, i.e. had the Pk phenotype. Sequencing revealed homozygosity for a nonsense mutation, c.420T>G, in B3GALNT1. This substitution introduces a premature stop codon, p.Tyr140Ter, which is predicted to abolish enzymatic activity. Screening of 384 Thai donors indicated an allele frequency of 0·13%. CONCLUSION: We describe a novel nonsense mutation (c.420T>G) in B3GALNT1 (GLOB*01N·13), which was added to the 12 alleles already known in the GLOB system.

Thorough analysis of unorthodox ABO deletions called by the 1000 Genomes project.

BACKGROUND AND OBJECTIVES: ABO remains the clinically most important blood group system, but despite earlier extensive research, significant findings are still being made. The vast majority of catalogued ABO null alleles are based on the c.261delG polymorphism. Apart from c.802G>A, other mechanisms for O alleles are rare. While analysing the data set from the 1000 Genomes (1000G) project, we encountered two previously uncharacterized deletions, which needed further exploration. MATERIALS AND METHODS: The Erythrogene database, complemented with bioinformatics software, was used to analyse ABO in 2504 individuals from 1000G. DNA samples from selected 1000G donors and African blood donors were examined by allele-specific PCR and Sanger sequencing to characterize predicted deletions. RESULTS: A 5821-bp deletion encompassing exons 5-7 was called in twenty 1000G individuals, predominantly Africans. This allele was confirmed and its exact deletion point defined by bioinformatic analyses and in vitro experiments. A PCR assay was developed, and screening of African samples revealed three donors heterozygous for this deletion, which was thereby phenotypically established as an O allele. Analysis of upstream genetic markers indicated an ancestral origin from ABO*O.01.02. We estimate this deletion as the 3rd most common mechanism behind O alleles. A 24-bp deletion was called in nine individuals and showed greater diversity regarding ethnic distribution and allelic background. It could neither be confirmed by in silico nor in vitro experiments. CONCLUSION: A previously uncharacterized ABO deletion among Africans was comprehensively mapped and a genotyping strategy devised. The false prediction of another deletion emphasizes the need for cautious interpretation of NGS data and calls for strict validation routines.

A and B antigen levels acquired by group O donor-derived erythrocytes following ABO-non-identical transfusion or minor ABO-incompatible haematopoietic stem cell transplantation.

BACKGROUND AND OBJECTIVES: ABO-incompatible haematopoietic stem cell transplantation (HSCT) presents a challenge to blood component transfusion. The aim of this study was to investigate the weak blood group A or B antigen expression by donor-derived group O red blood cells (RBC) observed following transfusion or minor ABO-incompatible HSCT. In addition, in vitro experiments were performed to elucidate possible mechanisms underlying this phenomenon. MATERIALS AND METHODS: A sensitive flow cytometry assay for the semi-quantification of RBC A/B antigen levels was used to assess patient samples and evaluate in vitro experiments. RESULTS: Analysis of blood samples from patients, originally typed as A, B and AB but recently transplanted or transfused with cells from group O donors, revealed the A antigen expression on donor-derived RBC, ranging from very low levels in non-secretor individuals to almost subgroup Ax -like profiles in group A secretors. The B antigen expression was less readily detectable. In vitro experiments, in which group O donor RBC were incubated with (i) group A/B secretor/non-secretor donor plasma or (ii) group A/B donor RBC in the absence of plasma, supported the proposed adsorption of A/B antigen-bearing glycolipids from secretor plasma but also indicated a secretor-independent mechanism for A/B antigen acquisition as well as direct cell-to-cell transfer of ABO antigens. CONCLUSION: The in vivo conversion of donor-derived blood group O RBC to ABO subgroup-like RBC after transfusion or minor ABO-incompatible HSCT raises the question of appropriate component selection. Based on these data, AB plasma should be transfused following ABO-incompatible HSCT.

International society of blood transfusion working party on red cell immunogenetics and terminology: report of the Seoul and London meetings.

The Working Party has met twice since the last report: in Seoul, South Korea 2014, and in London, UK 2015, both in association with the International Society of Blood Transfusion (ISBT) Congress. As in previous meetings, matters pertaining to blood group antigen nomenclature were discussed. Eleven new blood group antigens were added to seven blood group systems. This brings the current total of blood group antigens recognized by the ISBT to 346, of which 308 are clustered within 36 blood groups systems. The remaining 38 antigens are currently unassigned to a known blood group system.

International Society of Blood Transfusion Working Party on red cell immunogenetics and blood group terminology: Cancun report (2012).

The International Society of Blood Transfusion Working Party on red cell immunogenetics and blood group terminology convened during the International congress in Cancun, July 2012. This report details the newly identified antigens in existing blood group systems and presents three new blood group systems.

Six iterative reconstruction algorithms in brain CT: a phantom study on image quality at different radiation dose levels.

OBJECTIVE: To evaluate the image quality produced by six different iterative reconstruction (IR) algorithms in four CT systems in the setting of brain CT, using different radiation dose levels and iterative image optimisation levels. METHODS: An image quality phantom, supplied with a bone mimicking annulus, was examined using four CT systems from different vendors and four radiation dose levels. Acquisitions were reconstructed using conventional filtered back-projection (FBP), three levels of statistical IR and, when available, a model-based IR algorithm. The evaluated image quality parameters were CT numbers, uniformity, noise, noise-power spectra, low-contrast resolution and spatial resolution. RESULTS: Compared with FBP, noise reduction was achieved by all six IR algorithms at all radiation dose levels, with further improvement seen at higher IR levels. Noise-power spectra revealed changes in noise distribution relative to the FBP for most statistical IR algorithms, especially the two model-based IR algorithms. Compared with FBP, variable degrees of improvements were seen in both objective and subjective low-contrast resolutions for all IR algorithms. Spatial resolution was improved with both model-based IR algorithms and one of the statistical IR algorithms. CONCLUSION: The four statistical IR algorithms evaluated in the study all improved the general image quality compared with FBP, with improvement seen for most or all evaluated quality criteria. Further improvement was achieved with one of the model-based IR algorithms. ADVANCES IN KNOWLEDGE: The six evaluated IR algorithms all improve the image quality in brain CT but show different strengths and weaknesses.

An update on the GLOB blood group system and collection.

The P blood group antigen of the GLOB system is a glycolipid structure, also known as globoside, on the red blood cells (RBCs) of almost all individuals worldwide. The P antigen is intimately related to the Pk and NOR antigens discussed in the review about the P1PK blood group system. Naturally occurring anti-P is present in the serum of individuals with the rare globoside-deficient phenotypes p, P1k, and P2k and has been implicated in hemolytic transfusion reactions as well as unfavorable outcomes of pregnancy. The molecular genetic basis of globoside deficiency is absence of functional P synthase as a result of mutations at the B3GALNT1 locus. Other related glycolipid structures, the LKE and PX2 antigens, remain in the GLOB blood group collection pending further evidence about the genes and gene products responsible for their synthesis.

P1PK: the blood group system that changed its name and expanded.

The antigens in the P1PK blood group system are carried on glycosphingolipids. The system currently includes three different antigens, P1, Pk, and NOR. The P1 antigen was disovered in 1927 by Landsteiner and Levine, and Pk and NOR were described in 1951 and 1982, respectively. As in the ABO system, naturally occurring antibodies of the immunoglobulin (Ig) M or IgG class, against the missing carbohydrate structures, can be present in the sera of people lacking the corresponding antigen. Anti-P1 is generally a weak and cold-reactive antibody not implicated in hemolytic transfusion reaction (HTR) or hemolytic disease of the fetus and newborn while Pk antibodies can cause HTR, and anti-NOR is regarded as a polyagglutinin. A higher frequency of miscarriage is seen in women with the rare phenotypes p, P1k, and P2k. Furthermore, the Pk and P1 antigens have wide tissue distributions and can act as host receptors for various pathogens and toxins. Why p individuals lack not only Pk and P expression but also P1 has been a longstanding enigma. Recently, it was shown that the same A4GALT-encoded galactosyltransferase synthesizes both the P1 and Pk antigens and that a polymorphism in a new exon in this gene predicts the P1 and P2 phenotypes.

A novel B(weak) hybrid allele lacks three enhancer repeats but generates normal ABO transcript levels.

BACKGROUND AND OBJECTIVES: Weak expression of A/B histo-blood group antigens is often explained by single nucleotide substitutions at the ABO locus. However, hybrid alleles containing segments from different ABO alleles can result in unexpected phenotypes and may complicate genotype analysis. We investigated the basis of weak B phenotype in a referred sample. MATERIALS AND METHODS: A healthy young woman was serologically phenotyped as AB(weak) and RBCs were characterized by flow cytometry. All seven ABO exons, five introns plus the 5'-region including the CCAAT-binding factor/Nuclear Factor Y (CBF/NF-Y) binding enhancer were sequenced. ABO transcript levels were measured in fresh peripheral blood samples. Expression of B antigen was semiquantified following transfection of HeLa cells. RESULTS: A new B(weak) allele with 53G>T resulted in a characteristic pattern of moderately weakened B antigen expression on RBCs. Its sequence revealed a novel hybrid between O(2) [O03] and B [B101] alleles with a crossingover region in intron 4 as defined by allele-specific polymorphisms. B transcript levels were similar to normal controls despite the O(2) -related single CBF/NF-Y-binding 43-bp motif in the enhancer region. Expression of the glycosyltransferase including the O(2) -specific Arg18Leu substitution resulted in a slight decrease in B-antigen-positive cells. CONCLUSION: We describe here the first hybrid between an O(2) and a B allele and characterized the associated decrease in B antigen expression. Although it lacks three enhancer repeat units compared to common B alleles, the resulting transcript level was unaltered. This study challenges previous suggestions that the number of 43-bp motifs in the ABO enhancer determines transcription rates in erythroid cells.

Report of the fourth International Workshop on molecular blood group genotyping.

The fourth International Society of Blood Transfusion (ISBT) workshop on molecular blood group genotyping was held in 2010, with a feedback meeting at the ISBT Congress in Berlin, Germany. Fifty laboratories participated, 17 more than in 2008. Six samples were distributed. Samples 1-3 were DNA samples for all red cell blood group tests available to the participants. Of the 46 laboratories that tested these samples, 37 obtained completely correct results, although the extent of testing varied considerably. Sample 4, also a DNA sample, was an Rh problem in which RHDΨ and RHCE*ceCF were present, but the participants were only informed that the donor's red cells typed as positive with some monoclonal anti-D. Of the 42 laboratories that participated in this exercise, seven performed the sequencing necessary to obtain the correct result. Samples 5 and 6 were plasma samples from RhD-negative pregnant women, for foetal RhD testing. These were sent to 25 laboratories, and two incorrect results were reported. Overall, the level of accuracy was about equal to that of the previous workshop. The main conclusion for the last two workshops can be reiterated: with greater care and attention to detail, very high standards could be set for molecular blood group genotyping.

KEL*02 alleles with alterations in and around exon 8 in individuals with apparent KEL:1,-2 phenotypes.

BACKGROUND AND OBJECTIVES: Antibodies to antigens in the Kell blood group system, especially anti-KEL1, are involved in both haemolytic disease of the newborn and foetus and haemolytic transfusion reactions. Correct typing results are important and discrepancies between serologic and genetic typing must be resolved. Here, we describe the investigation of three healthy individuals who were initially phenotyped as KEL:1,-2. MATERIALS AND METHODS: Antigen typing was performed by standard serological techniques and by flow cytometric analysis. The KEL*01/02 polymorphism was tested by an allele-discrimination TaqMan assay as well as by PCR with allele-specific primers and PCR-RFLP. DNA sequencing of the KEL coding region was also performed. RESULTS: Two KEL*02N alleles with mutated splice sites around exon 8 were identified: intron 7 -1g>c (novel) and intron 8 +1g>t (previously reported in one case of K(0)). In the third sample, a missense mutation in exon 8, 787G>A (novel) predicting Gly263Arg, was detected on a KEL*02 allele and associated with dramatically weakened KEL2 antigen expression. CONCLUSION: Resolution of discrepant phenotype/genotype results identified silencing mutations in or around exon 8. A combination of molecular and serologic methods has the potential to improve the quality of test results and was required to ensure both the accurate KEL2 antigen status and KEL*01 zygosity of these individuals.

A phantom study showing the importance of compression in conventional diagnostic X-ray examinations.

Compression was earlier an important and well-managed part of the clinical routine, but during the past 15-20 y the use has diminished, except for mammography, where it is a prerequisite for having images of high quality and low radiation dose. According to national and European regulations and recommendations, it is important to apply the compression technique to obtain the optimal image quality and radiation dose in common conventional diagnostic examinations. Current experience of compression technique is, however, based on studies carried out a long time ago and with analogue imaging techniques. An anthropomorphic phantom was used to show the importance of compression in conventional X-ray examinations. The patient thicknesses on volunteers with and without compression was measured. This measurement was done to investigate compression potential on patients and to select suitable phantom thicknesses. The X-ray examinations that were included in the study were abdomen overview, lumbar spine and the pelvis. The results from the phantom study showed a large dependency of the kerma-area product value on the phantom thickness. The phantom study suggests that there is a potential for significant reduction of radiation dose to the patient by using compression also with modern X-ray techniques. A dose reduction of up to 50 % or even more may be obtained.

The ABO blood group system revisited: a review and update.

The antigens of the ABO system were the first to be recognized as blood groups and actually the first human genetic markers known. Their presence and the realization of naturally occurring antibodies to those antigens lacking from the cells made sense of the erratic failure of blood transfusion hitherto and opened up the possibility of a safe treatment practice in life-threatening blood loss. Although initially apparently simple, the ABO system has come to grow in complexity over the years. The mass of knowledge relating to carbohydrate chemistry, enzymology, molecular genetics, and structural and evolutionary biology is now enormous thanks to more than a century of research using ABO as a principal model. This has provided us with data to form a solid platform of evidence-based transfusion and transplantation medicine used every day in laboratories and clinics around the globe. This review aims to summarize key findings and recent progress made toward further understanding of this surprisingly polymorphic system.

Report of the third international workshop on molecular blood group genotyping.

The Third International Society of Blood Transfusion Workshop on Molecular Blood Group Genotyping was held in 2008, with a feedback meeting at the International Society of Blood Transfusion Congress in Macao SAR, China. Thirty-three laboratories participated, eight less than in 2006. Six samples were distributed: sample 1 representing DNA from a sample referred because of abnormal serological results in D testing; samples 2 and 3 from transfusion-dependent patients for testing for all clinically important polymorphisms; sample 4 a mixture of two DNA samples designed to simulate a chimera, referred because of abnormal serological results in donor testing; and samples 5 and 6 plasma samples from RhD-negative pregnant women, for fetal RhD testing (only tested by 17 laboratories). For samples 1-3, 24 of 33 laboratories obtained completely correct results. For sample 4, the ability to detect the minority DNA population was partly dependent on method. Of the 17 laboratories that received samples 5 and 6, 13 reported correct results on both samples. Overall a small improvement from previous workshops was noted, but there is still room for improvement. The main conclusion for the 2006 workshop can be reiterated: with greater care and attention to detail, very high standards could be set for molecular blood group genotyping.