Blood Group Polymorphisms Research Articles

The Presence of FYAnull Alele of Duffy Blood Group System in Blood Donors and Individuals from a Malarial Endemic Region of Brazil.

BACKGROUND: Malaria is a virulent disease caused by the Plasmodium parasite. Innate resistance to malaria infections in humans is conferred by various blood group polymorphisms. The Duffy blood group system consists of Fya and Fyb antigens which are encoded by codominant alleles FYA and FYB. Four phenotypes are defined: Fy(a+b+), Fy(a+b−), Fy(a−b+) and Fy(a−b−). Erythrocytes of Duffy-negative individuals are resistant to invasion by P. vivax. In Blacks the Fy(a−b−) phenotype is associated with a single point mutation (-33T-C) in the GATA-1 binding motif for the erythroid promoter of FYB.STUDY DESIGN AND METHODS: We investigated the phenotypes and the genotypes of Duffy blood group system of 250 individuals living in a malarial endemic region (MER) in the state of Amazon (Brazil), and of 199 blood donors (BD) from a non-endemic region. The phenotyping was done by agglutination gel tests (DiaMed-Latino América) using anti-Fya and anti-Fyb reagents. The molecular analysis for FYA, FYB, FYBES (GATA box mutation nt -33T-C), and FYBWeak (mutations 265 C-T, and 298 G-A) alleles, were performed by PCR-RFLP. The PCR products were digested by Ban I for FYA and FYB identifications; by Sty I for GATA box mutation; Acy I and Mwo I for 265 C-T and 298 G-A mutations, respectively. Some samples that showed discrepancy between the phenotype and genotype results were examined by sequence analysis using the ABI PrismâBig Dyeä Terminator Cycle Sequencing Ready Reaction Kit” (Perkin Elmer), and the interpretation by the software ABI PRISMä 377 DNA Sequencer”, 3.3 version (Perkin Elmer).RESULTS: We found that 34/250 (13.6%) of 250 persons living in the MER and 37/199 (18.6%) of BD had phenotype and genotype discrepant results [Fy(a+b−) FYA/FYB]. In addition, we found that 16/34 (47%) of people living in the MER, and 4/37 (10.8%) of BD did not present the -33T-C mutation, the 265 C-T, or the 298 G-A mutations. The sequence analysis of 2 samples from persons from MER indicated the presence of -33T-C mutation in the FYA allele in one individual (1 FYA/FYB and W/M; FYA/FYB and M/M). Additionally, we detected that 18/34 (53%) of people living in the MER, and 33/37 (89.2%) of BD presented the -33T-C mutation. The sequence analysis of 5 samples indicated the presence of -33T-C mutation in the FYA allele in 4 cases [2 persons from MER and 2 from BD (FYA/FYB e M/M)].CONCLUSION: Recently the mutation responsible for erythrocyte Duffy antigen-negativity [Fy(a−b−)] was demonstrated in FYA allele in a malarial endemic region of Papua New Guinea. The present data demonstrated the presence of the FYAnull allele not only in persons living in a malarial endemic region but also in Brazilian blood donors from non-endemic areas. In contrast with that which happens with the FYB allele, our results indicated that the presence of the -33T-C mutation in the FYA allele does not abolish the expression of the Fya antigen in the erythrocyte.

Increased Von Willebrand Factor Antigen Clearance in Type 1 Von Willebrand Disease: Investigation of Relationship with ABO Blood Group and Tyr1584Cys Polymorphism.

Type 1 von Willebrand disease (VWD) is a heterogeneous bleeding disorder in which genetic modifying factors, including ABO blood group, contribute towards the variability in von Willebrand factor (VWF) levels. Recent findings have reported an increased incidence of the Tyr1584Cys polymorphism in the VWF-A2 domain of patients with type 1 VWD. Presence of Cys1584 has been shown to cause increased intracellular retention of VWF, as well as lead to increased susceptibility of VWF to proteolysis by the metalloprotease ADAMTS13. An increased susceptibility to proteolysis by ADAMTS13 has also been demonstrated in the VWF of blood group O individuals. We have investigated the relationship between increased VWF antigen (VWF:Ag) clearance, ABO blood group and the presence of the Tyr1584Cys polymorphism in a group of patients with type 1 VWD. The VWF:Ag half-life (VWF:Ag t1/2) was prospectively evaluated in 45 patients with type 1 VWD, which included three families of two or more first-degree relatives. Median VWF:Ag level was 36.0iu/dl (range 4–50iu/dl); median VWF ristocetin cofactor activity (VWF:RCo) level was 35.0iu/dl (range 3–52iu/dl) and VWF:RCo/VWF:Ag ratio 0.97 (range 0.70–1.37). A normal multimeric pattern was demonstrated in all patients. 25 (55.5%) and 20 (44.5%) of the patients were of blood groups O and A respectively. The control group comprised eight patients with haemophilia A. VWF:Ag levels were measured over a 6 hour period following the administration of intravenous DDAVP. VWF:Ag t1/2 was calculated using the formula: C(t)=C0e−k.t, where C(t)=plasma [VWF:Ag] as a function of time; C0=[VWF:Ag] at time zero; e=base for natural logarithms; k=first order rate constant for the elimination phase (ß phase); t=time. The median value of the VWF:Ag t1/2 in the VWD group was 4.1 hours (95% C.I. 3.2–4.9h) and in the haemophilia A group 9.5 hours (95% C.I. 5.3–19h). This represents a significant difference in VWF:Ag t1/2 between the two groups (p<0.05). However, within the VWD patient group, there was no significant difference between the median VWF:Ag t1/2 values of patients of blood group O and those of blood group A (p>0.05). Within the three families, two affected family members of the same ABO blood group were studied and a concordant reduction in the VWF:Ag t1/2s was found in these subjects. To date, 24 of the VWD patients have been genotyped for the A/G polymorphism at nucleotide 24/1282 in the VWF gene, encoding a Tyr1584Cys polymorphism. The heterozygous presence of the G allele encoding Cys 1282 was demonstrated in one patient. The frequency of this polymorphism in normal and type 1 VWD individuals has been reported to be ~1% and 14% respectively. The median VWF:Ag t1/2 value of the 23 homozygous Tyr1584 patients was 3.4 hours (95% C.I. 3.2–4.8h), representative of the whole VWD group. The VWF:Ag t1/2 in the heterozygous patient was 4.8 hours. The finding of increased plasma VWF:Ag clearance as reflected by a reduction in VWF:Ag half-life in a significant number of patients with type 1 VWD, suggests that increased VWF:Ag clearance may be a contributory factor in the aetiology of type 1 VWD. However, this study suggests there is no relationship between increased VWF:Ag clearance and ABO blood group. Furthermore, the Tyr1584Cys polymorphism is not a major determinant of VWF:Ag clearance within this group of type 1 VWD patients.

Fetal blood group genotyping from DNA from maternal plasma: an important advance in the management and prevention of haemolytic disease of the fetus and newborn

The cloning of blood group genes and subsequent identification of the molecular bases of blood group polymorphisms has made it possible to predict blood group phenotypes from DNA with a reasonable degree of accuracy. The major application of this technology, which has now become the standard of care, is the determination of a fetal RHD genotype in women with anti-D, to assess whether the fetus is at risk of haemolytic disease of the fetus and newborn (HDFN). Initially, the procurement of fetal DNA required the invasive procedures of amniocentesis or chorionic villus sampling. Since the discovery of fetal DNA in maternal plasma in 1997, the technology of detecting an RHD gene in this very small quantity of fetal DNA has developed rapidly, so that non-invasive fetal D typing can now be provided as a diagnostic service for D-negative pregnant women with anti-D. Within a few years, it is probable that fetuses of all D-negative pregnant women will be tested for RHD, to establish whether the mother requires antenatal anti-D immunoglobulin prophylaxis.

AGT and RH blood group polymorphisms affect blood pressure and lipids in Afro-Caribbeans.

Population blood pressure variation is most likely due to multiple genes. This is likely the reason why monogenic testing with the angiotensinogen (AGT) gene polymorphisms on chromosome 1 (1q42-43), especially M235T, has met with negative results, especially in those of African descent. The RH blood group system, also on chromosome 1 (1 p36.2-34), has likewise been associated with blood pressure variation in African-Americans and with the rise in blood pressure with age in whites. Using a random sample of the population, we investigated the combined effects of single and combined variation of the AGTN M235T and RH genotypes on blood pressure, lipids, and lipoprotein concentrations in Afro-Caribbeans aged 18-60 years from the island nation of Dominica. In monogenic analysis, AGT M235T was not associated with blood pressure. However, it was associated with HDL (MM 42+/-23, MT 44+/-12, TT 52+/-14 (P=0.002)). RH genotype was significantly associated with systolic blood pressure (P=0.006) and Apo-A (P=0.003). These effects remained after adjustment for age, gender, weight, and BMI. In the polygenetic analysis, AGT M235T and RH were significantly associated with systolic blood pressure (P=0.037; interaction effects, P=0.068). The association of the AGT M235T with blood pressure across RH blood group haplotypes was then tested. Of the five RH haplotypes available for analysis, the AGT M235T was significantly associated with blood pressure within the "D" haplotype (P=0.01). The RH blood group and gender were significantly associated with systolic blood pressure and Apo-A levels (P=0.005 and 0.012, respectively). All interactions were independent of age and weight. In conclusion, we demonstrate a significant association of AGT M235T with blood pressure and cholesterol metabolism in an Afro-Caribbean population in the "genetic context" of the RH blood group system. Further investigation of these interactions may help understand the effects of genetic factors on cardiovascular risk in African-derived and other populations.

Blood group genotyping

Accurate phenotyping of red blood cells (RBCs) can be difficult in transfusion-dependent patients such as those with thalassemia and sickle cells anemia because of the presence of previously transfused RBCs in the patient's circulation. Recently, the molecular basis associated with the expression of many blood group antigens was established. This allowed the development of a plethora of polymerase chain reaction-based tests for identification of the blood group antigens by testing DNA. The determination of blood group polymorphism at the genomic level facilitates the resolution of clinical problems that cannot be addressed by hemagglutination. They are useful to (a) determine antigen types for which currently available antibodies are weakly reactive; (b) type patients who have been recently transfused; (c) identify fetuses at risk for hemolytic disease of the newborn; and (d) to increase the reliability of repositories of antigen negative RBCs for transfusion. It is important to note that PCR based assays are prone to different types of errors that those observed with hemagglutination assays. For instance, contamination with amplified products may lead to false positive test results. In addition, the identification of a particular genotype does not necessarily mean that the antigen will be expressed on the RBC membrane.

Karl landsteiner and his major contributions to haematology

Karl landsteiner and his major contributions to haematology

ABO blood group in Amerindians from Brazilian Amazon

Background : The Parakanã is a group of Indians with cultural similarities to the extinct Tupi group. They are an isolated native population from East Brazilian Amazon. A number of different O alleles have been found at the blood group ABO locus in populations of several ethnic origins (Caucasians, Blacks, Amerindians). Aim : The present study describes the ABO blood group polymorphism gene of the Parakanã Indians. The Amerindian group was carefully selected for racial background. Subject and methods : The blood group polymorphism was analysed in genomic DNA from 62 Parakanã Indians. We determined the 261G deletion, the T646A and C771T mutations described in O 1variant and the G542A substitution, using PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism). Results : All Amerindians studied were homozygous for the 261G deletion. The frequencies of the T646A and C771T mutations in Parakanãs (0.65) were lower than that observed in Kayapo, Yanomama and Arara Indians (0.91) ( h 2 = 18.24; p-v < 0.001. The G542A substitution in Parakanãs was also lower (0.22) than in other tribes (0.42) ( h 2 = 9.73; p-v = 0.001). Conclusions : The different O alleles including the G542A mutation are not distributed homogeneously among all Amazonian Amerindians. Our results are in agreement with other genetic markers studied previously in Parakanã Indians, whose distinct genetic pattern differs from Europeans and even from other Amerindians.

Antigenic and Functional Properties of the Human Red Blood Cell Urea Transporter hUT-B1

The Kidd (JK) blood group locus encodes the urea transporter hUT-B1, which is expressed on human red blood cells and other tissues. The common JK*A/JK*B blood group polymorphism is caused by a single nucleotide transition G838A changing Asp-280 to Asn-280 on the polypeptide, and transfection of erythroleukemic K562 cells with hUT-B1 cDNAs carrying either the G838 or the A838 nucleotide substitutions resulted in the isolation of stable clones that expressed the Jk(a) or Jk(b) antigens, respectively, thus providing the first direct demonstration that the hUT-B1 gene encodes the Kidd blood group antigens. In addition, immunochemical analysis of red blood cells demonstrated that hUT-B1 also exhibits ABO determinants attached to the single N-linked sugar chain at Asn-211. Moreover, immunoadsorption studies, using inside-out and right-side-out red cell membrane vesicles as competing antigen, demonstrated that the C- and N-terminal ends of hUT-B1 are oriented intracellularly. Mutagenesis and functional studies by expression in Xenopus oocytes revealed that both cysteines Cys-25 and Cys-30 (but not alone) are essential for plasma membrane addressing. Conversely, the transport function was not affected by the JK*A/JK*B polymorphism, C-terminal deletion (residues 360-389), or mutation of the extracellular N-glycosylation consensus site and remains poorly para-chloromercuribenzene sulfonate (pCMBS)-sensitive. However, transport studies by stopped flow light scattering using Jk-K562 transfectants demonstrated that the hUT-B1-mediated urea transport is pCMBS-sensitive in an erythroid context, as reported previously for the transporter of human red blood cells. Mutagenesis analysis also indicated that Cys-151 and Cys-236, at least alone, are not involved in pCMBS inhibition. Altogether, these antigenic, topologic, and functional properties might have implications into the physiology of hUT-B1 and other members of the urea transporter family.

Molecular identification of Knops blood group polymorphisms found in long homologous region D of complement receptor 1: J.M. Moulds, P.A. Zimmerman, D.K. Doumbo, et al. Blood 97:2879–2885, 2001

Molecular identification of Knops blood group polymorphisms found in long homologous region D of complement receptor 1: J.M. Moulds, P.A. Zimmerman, D.K. Doumbo, et al. Blood 97:2879–2885, 2001

Genetic characterisation of the Uruguayan Creole horse and analysis of relationships among horse breeds

The genetic variability within the Uruguayan Creole horse and its relationship to a group of geographically or historically related breeds (Spanish Pure-bred, Barb, Quarter horse, Paso Fino, Peruvian Paso, Arabian and Thoroughbred horse), was evaluated using 25 loci (seven of blood groups, nine of protein polymorphisms and nine microsatellites) analyzed on a total of 145 Uruguayan Creole horses. In this study, blood group and protein polymorphism variants that are considered to be breed markers of Spanish Pure-bred and Barb horses were detected in the Creole breed. Conversely, some microsatellites and protein polymorphisms alleles were found uniquely in the Creole horse. American horse breeds together with Barb and Arabian horses clearly formed a separate cluster from the Spanish pure-bred and Thoroughbred breeds, as shown by anupgma dendrogram based on Nei's standard genetic distance. Data in this study provided evidence for considerable genetic variation within Uruguayan Creole horses and of a distinctive breed profile. Both traits were most likely inherited from the XVIth century Spanish horses, more closely related to Barb than to Spanish Pure-bred.

Immunization of transgenic mice for production of MoAbs directed at polymorphic blood group antigens.

Antibodies of human origin for blood typing are increasingly difficult to obtain, and, despite aggressive efforts, MoAbs with specificities to several blood group polymorphisms have eluded production. As an approach for the generation of MoAbs with defined specificities, the feasibility of immunizing mice that are transgenic for the target polymorphism, Fy(a)/Fy(b) of the Duffy blood group system, was tested with a source of the antithetical antigen. Nontransgenic mice were immunized with recombinant Fy(b), and transgenic mice expressing human Fy(b) were immunized with recombinant Fy(a). Immunization of the nontransgenic mice resulted in the production of MoAbs to the Duffy protein, but not to the Fy(a)/Fy(b) blood group polymorphism. However, immunization of the transgenic mice resulted in production of the first example of murine Fy(a) MoAb (MIMA-19). This antibody is being used to screen for Fy(a-) blood donors and has been evaluated by many laboratories in an international workshop. This approach provides an effective method for producing MoAbs with specificities to polymorphic epitopes. These MoAbs are needed in transfusion medicine to identify antigen-negative donors and to alleviate the critical shortage of blood bank typing reagents, which currently are available only from human-derived sources.

A molecular genetic analysis of the Kidd blood group polymorphism in Taiwanese, Thais, and Filipinos.

In this study, we molecularly detect the Jka/Jkb polymorphism in Taiwanese, Thais, and Filipinos with PCR-restriction enzyme digestion. We designed the primers with mutagenic bases to create two restriction enzyme cutting sites (EcoN I and Xba I) simultaneously to confirm the genetic polymorphism of Jka/Jkb. One hundred and seven unrelated Taiwanese, 107 Thais foreign laborers, and 102 Filipino foreign laborers in Taiwan were analyzed. The frequency of Jka and Jkb is 0.51 and 0.49 in Taiwanese, 0.44 and 0.56 in Thais, and 0.45 and 0.55 in Filipinos, respectively. The molecular analysis of Jka/Jkb polymorphism is simple, accurate and could confirm the uncertain results obtained by serological typing. The method could be used for predicting fetal Kidd genotype from amniocyte DNA, and to avoid the hemolytic disease of newborns caused by Jk antibody.

Genomic analysis of clinical samples with serologic ABO blood grouping discrepancies: identification of 15 novel A and B subgroup alleles

Since the cloning in 1990 of complementary DNA corresponding to messenger RNA transcribed at the blood group ABO locus, polymorphisms and phenotype-genotype correlations have been reported by several investigators. Exons 6 and 7, constituting 77% of the gene, have been analyzed previously in samples with variant phenotypes but for many subgroups the molecular basis remains unknown. This study analyzed 324 blood samples involved in ABO grouping discrepancies and determined their ABO genotype. Samples from individuals found to have known subgroup alleles (n = 53), acquired ABO phenotypes associated with different medical conditions (n = 65), probable chimerism (n = 3), and common red blood cell phenotypes (n = 109) were evaluated by ABO genotype screening only. Other samples (n = 94) from apparently healthy donors with weak expression of A or B antigens were considered potential subgroup samples without known molecular background. The full coding region (exons 1-7) and 2 proposed regulatory regions of the ABO gene were sequenced in selected A (n = 22) or B (n = 12) subgroup samples. Fifteen novel ABO subgroup alleles were identified, 2 of which are the first examples of mutations outside exon 7 associated with weak subgroups. Each allele was characterized by a missense or nonsense mutation for which screening by allele-specific primer polymerase chain reaction was performed. The novel mutations were encountered in 28 of the remaining 60 A and B subgroup samples but not among normal donors. As a result of this study, the number of definable alleles associated with weak ABO subgroups has increased from the 14 previously published to 29.

Molecular identification of Knops blood group polymorphisms found in long homologous region D of complement receptor 1

AbstractComplement receptor 1 (CR1) has been implicated in rosetting of uninfected red blood cells to Plasmodium falciparum–infected cells, and rosette formation is associated with severe malaria. The Knops blood group (KN) is located on CR1 and some of these antigens, ie, McCoy (McC) and Swain-Langley (Sla), show marked frequency differences between Caucasians and Africans. Thus, defining the molecular basis of these antigens may provide new insight into the mechanisms of P falciparummalaria. Monoclonal antibody epitope mapping and serologic inhibition studies using CR1 deletion constructs localized McC and Slato long homologous repeat D of CR1. Direct DNA sequencing of selected donors identified several single nucleotide polymorphisms in exon 29 coding for complement control protein modules 24 and 25. Two of these appeared to be blood group specific: McC associated with K1590E and Sla with R1601G. These associations were confirmed by inhibition studies using allele-specific mutants. A sequence-specific oligonucleotide probe hybridization assay was developed to genotype several African populations and perform family inheritance studies. Concordance between the 1590 mutation and McC was 94%; that between Sla and 1601 was 88%. All but 2 samples exhibiting discrepancies between the genotype and phenotype were found to be due to low red cell CR1 copy numbers, low or absent expression of some alleles, or heterozygosity combined with low normal levels of CR1. These data further explain the variability observed in previous serologic studies of CR1 and show that DNA and protein-based genetic studies will be needed to clarify the role of the KN antigens in malaria.

A DNA-based immunization protocol to produce monoclonal antibodies to blood group antigens.

A major challenge facing transfusion medicine is the establishment of immunological methods to produce specific and avid blood group typing reagents to the many polymorphic blood group antigens. This is especially true when sources of human antibody are limited. Based on the knowledge that inoculation with plasmid DNA can induce a humoral response in the host animal, we inoculated mice with plasmid DNA followed by a single boost injection with plasmid-transfected cells that have a high level of expression of the same target protein. Using this method, several hybridoma clones that produced strongly reactive antibodies specific for the Kell polymorphic antigens (anti-K, anti-k, anti-Kp(a)) were isolated. The monoclonal antibodies that were produced with this method have potential clinical utility for identifying a patient's blood type and for screening for antigen-negative donor blood.

An amino acid substitution in the putative second extracellular loop of RBC band 3 accounts for the Froese blood group polymorphism.

The low incidence RBC antigen Fr(a) has been excluded from 17 of the 25 established blood group systems. Previous genetic analysis assigned the gene controlling Fr(a) expression to the same chromosomal region as the solute carrier family 4, anion exchanger member 1 gene (SLC4A1). Because SLC4A1 encodes RBC band 3 and controls the expression of Diego blood group system antigens, the possible relationship of Fr(a) to the Diego blood group system was investigated by molecular analysis of SLC4A1. Blood samples were obtained from the members of two unrelated Mennonite kindreds segregating for Fr(a). DNA was extracted, amplified by PCR using intronic primer sets flanking exons 11-20 of SLC4A1, and screened by single-strand conformation polymorphism (SSCP) analysis. Those exons displaying SSCPs were subjected to DNA sequence analysis. An exon 13 SSCP mobility shift was observed in the DNA from all Fr(a+) persons that was not seen in the DNA from Fr(a-) family members or control subjects. Linkage between the exon 13 SSCP and FR:(a) was established, with peak lods = 3.62 at theta = 0.00 for combined paternal and maternal meioses. DNA sequencing revealed a GAG --> AAG mutation that underlies a Glu480Lys substitution in RBC band 3. A point mutation in exon 13 of SLC4A1 accounting for a Glu480Lys substitution in band 3 controls Fr(a) expression. On the basis of these our results, the International Society of Blood Transfusion Working Party on Terminology for Red Cell Surface Antigens has assigned Fr(a) to the Diego blood group system, with the designation DI20.

Quantitative analysis of XG blood group and CD99 antigens on human red cells.

CD99, the product of the MIC2 gene, exhibits an erythroid-specific quantitative polymorphism co-regulated with the Xga blood group polymorphism. The co-expression of X-linked MIC2 and XG genes is presumably controlled at the transcriptional level by a single XGR locus in the pseudoautosomal region of sexual chromosomes. This locus is composed of two alleles, XGR(low) and XGR(high), which determine low or high CD99 levels (CD99-L, CD99-H) and the Xg(a-)/ Xg(a+) status. To test this hypothesis, the phenotypic relationship between Xga and CD99 antigens on human RBCs was investigated by quantitative flow cytometry using NBL-1 (anti-Xga) and 12E7 (anti-CD99) monoclonal antibodies and semi-quantitative estimate of membrane proteins and RNA by Western blot and Northern blot, respectively. The antibody binding capacity of RBCs, which is an estimation of the antigen density, was determined for 118 blood donors including 60 males and 58 females. Xg(a+) RBCs, which all belong to the group of CD99-H expressors, carry 159+/-13 and 960+/-50 copies of Xga and CD99 molecules/cell, respectively. Xg(a-) RBCs have no Xga antigen, but are subdivided into CD99-H (all male) and CD99-L expressors carrying 747+/-28 and 200+/-22 CD99 copies/cell, respectively, with identical CD99 levels between CD99-L males and females. However, among males, the CD99 expression was higher in Xg(a+) than in Xg(a-)/CD99-H individuals (P<0.01). In addition, CD99-H expressors in Xg(a+) males could be clearly subdivided into two categories, high and super high expressors, which are presumably heterozygous and homozygous for the XGR(high) allele, which fits the above hypothesis. This was not the case for Xg(a+) females where CD99-H subcategories were not found. Quantitative differences were confirmed by Western blot analysis of red cell membrane preparations from individuals of different Xga and CD99 phenotypes and by Northern blot analysis showing that the reticulocytes from CD99-L individuals expressed a reduced level of MIC2 transcripts compared to CD99-H donors. These findings further support the hypothesis of a single genetic control of CD99 and Xga expression by the XGR locus.

Molecular biology and genetics of the Rh blood group system

The Rh (Rhesus) blood group system is the most complex of the known human blood group polymorphisms. The expression of its antigens is controlled by a two-component genetic system consisting of RH and RHAG loci, which encode Rh30 polypeptides and Rh50 glycoprotein, respectively. Over the past decade, there has been a rapid advance in knowledge of the biochemistry, molecular biology, and genetics of the Rh genes and proteins. The primary structures of D and CcEe antigens have become well understood and the molecular genetic basis of a vast array of phenotype polymorphisms has been delineated. The identification of various molecular defects associated with Rh deficiency syndrome clarifies the nature of the amorph, suppressor, and modifier genes. The observed mutation spectrum defines a basic set of components essential for Rh complex assembly in the erythrocyte membrane. The resulting molecular information, combined with new experimental tools, is helping to dissect the fine structure of Rh antigens in terms of epitope mapping. The discovery of novel Rh homologs in primitive organisms and in nonerythroid tissues opens new avenues of research beyond the scope of erythrocytes and Rh antigens. This review provides an update on the Rh family in antigen expression, phenotype diversity, and disease association.

Unravelling the biochemical basis of blood group ABO and Lewis antigenic specificity.

The ABO blood-group polymorphism is still the most clinically important system in blood transfusion practice. The groups were discovered in 1900 and the genes at the ABO locus were cloned nearly a century later in 1990. To enable this goal to be reached intensive studies were carried out in the intervening years on the serology, genetics, inheritance and biochemistry of the antigens belonging to this system. This article describes biochemical genetic investigations on ABO and the related Lewis antigens starting from the time in the 1940s when serological and classical genetical studies had established the immunological basis and mode of inheritance of the antigens but practically nothing was known about their chemical structure. Essential steps were the definition of H as the product of a genetic system Hh independent of ABO, and the establishment of the precursor-product relationship of H to A and B antigens. Indirect methods gave first indications that the specificity of antigens resided in carbohydrate and revealed the immunodominant sugars in the antigenic structures. Subsequently chemical fragmentation procedures enabled the complete determinant structures to be established. Degradation experiments with glycosidases revealed how loss of one specificity by the removal of a single sugar unit exposed a new specificity and suggested that biosynthesis proceeded by a reversal of this process whereby the oligosaccharide structures were built up by the sequential addition of sugar units. Hence, the primary blood-group gene products were predicted to be glycosyltransferase enzymes that added the last sugar to complete the determinant structures. Identification of these enzymes gave new genetic markers and eventually purification of the blood-group A-gene encoded N-acetylgalactosaminyltransferase gave a probe for cloning the ABO locus. Blood-group ABO genotyping by DNA methods has now become a practical possibility.

Emergence of FY*A(null) in a Plasmodium vivax-endemic region of Papua New Guinea.

In Papua New Guinea (PNG), numerous blood group polymorphisms and hemoglobinopathies characterize the human population. Human genetic polymorphisms of this nature are common in malarious regions, and all four human malaria parasites are holoendemic below 1500 meters in PNG. At this elevation, a prominent condition characterizing Melanesians is alpha(+)-thalassemia. Interestingly, recent epidemiological surveys have demonstrated that alpha(+)-thalassemia is associated with increased susceptibility to uncomplicated malaria among young children. It is further proposed that alpha(+)-thalassemia may facilitate so-called "benign" Plasmodium vivax infection to act later in life as a "natural vaccine" against severe Plasmodium falciparum malaria. Here, in a P. vivax-endemic region of PNG where the resident Abelam-speaking population is characterized by a frequency of alpha(+)-thalassemia >/=0.98, we have discovered the mutation responsible for erythrocyte Duffy antigen-negativity (Fy[a-b-]) on the FY*A allele. In this study population there were 23 heterozygous and no homozygous individuals bearing this new allele (allele frequency, 23/1062 = 0.022). Flow cytometric analysis illustrated a 2-fold difference in erythroid-specific Fy-antigen expression between heterozygous (FY*A/FY*A(null)) and homozygous (FY*A/FY*A) individuals, suggesting a gene-dosage effect. In further comparisons, we observed a higher prevalence of P. vivax infection in FY*A/FY*A (83/508 = 0.163) compared with FY*A/FY*A(null) (2/23 = 0.087) individuals (odds ratio = 2.05, 95% confidence interval = 0.47-8.91). Emergence of FY*A(null) in this population suggests that P. vivax is involved in selection of this erythroid polymorphism. This mutation would ultimately compromise alpha(+)-thalassemia/P. vivax-mediated protection against severe P. falciparum malaria.