Blood Groups: More than Inheritance of Antigenic Substances - Susceptibility to Some Diseases

*Williams Bitty Azachi and Kuschak Mathias Dakop*

## **Abstract**

Blood group antigens represent polymorphic traits inherited among individuals and populations. The objective of this chapter is to review articles that have reported; the association between blood group antigens and susceptibility to some diseases. Findings showed that O blood group had a greater frequency of severe infections such as E coli, cholera and blood group A was associated with incidence of smallpox and some bacterial infections. These are principally based on presence or absence of "H-like" and "A and B-like" antigens markers. Antigens A, B and H are connected to N-glycans of vWF and reduces the half-life of the protein (10 hours) for group O while non-O groups, 25 hours. The loss of A, B, and H antigens as malignancy progresses was linked to potential metastasis. Similarly, some tumors have A or A-like antigens this explains the propensity of group A to develop tumors. Blood type incompatibility between mother and foetus sensitizes the mother to develop alloantibodies that could potentially cause death of the foetus in utero, a condition known hydrops. Reviewed articles have reported close link between blood group antigens and susceptibility diseases. More studies are required to rationalize the mechanism associated to this.

**Keywords:** blood group antigen, susceptible, disease

## **1. Introduction**

Human blood groups since discovery in 1901 by Landsteiner have been widely studied and characterized. A total of 34 blood group systems have been recognized and documented by the International Society for Blood Transfusion (ISBT) [1, 2]. Each system is a series of red cell antigens, determined either by a single genetic locus or very closely linked loci. Alternative forms of a gene coding for red cell antigens at a particular locus are called alleles and individuals may inherited identical or non-identical alleles [3].

Many blood groups are receptors for toxins, parasites, and bacteria, where they can facilitate colonization or invasion or evade host clearance mechanisms [4]. Associations between the blood type and disease have been studied since the early 1900s when researchers determined that antibodies and antigens are inherited. However, due to lack of antigens of some blood groups, there have been some contentious issues with the association between the ABO blood group and vulnerability to certain infectious and non-infectious diseases [5].



#### **Table 1.**

*Blood group systems recognized by the International Society of Blood Transfusion.*

Fung et al. [1] (**Table 1**) gives blood group antigens characterized by ISBT.

## **2. ABO antigens linked to some diseases**

#### **2.1 Infectious disease**

ABO gene products have been associated with some diseases [6–8]. The human body defense integrity against certain infections is characteristic of the presence or absence of blood group antigens and their corresponding antibodies. More so,

#### *Blood Groups: More than Inheritance of Antigenic Substances - Susceptibility to Some Diseases DOI: http://dx.doi.org/10.5772/intechopen.104593*

data have shown that peptic ulceration was the first proven association to blood group gene products [9, 10]. *H pylori* is now known to be a causative agent leading to peptic ulceration and gastric cancer. *H pylori* has established colonies in the stomach of approximately one-half the world's population [11].

Red blood cell surface markers act as receptors for attachment to infectious agents and result in vulnerability difference among individuals with diverse receptor profile [12]. Some pathogens share genetic properties with their host. The relationship between ABO antigens and infections as vibrio cholera was discovered by early studies [12]. Major variations in ABO groups in the world were due to H-like antigen on the bacterium (*Vibrio cholera*) and an A-like antigen on Small pox virus [13]. This confers resistance status to people who make corresponding antibodies to H and A antigens [8]. Once a person gets infected with Cholera (Vibrio cholera strain O 1, E 1, Tor and O 139), the O blood types have a greater frequency of severe infections than the non-O blood types. Increased incidence of cholera is strongly associated to blood group O whereas, blood group A is strongly associated to smallpox, Pseudomoniasis, gonorrhea, tuberculosis and *streptococcus pneumoniae*, *Escherichia coli* and Salmonellosis. Blood type AB is linked to increased incidence of small pox, E. coli and Salmonellosis. Principally, these are associated to the presence of H antigen on group O and anti-H on group A and B [8].

The GI expresses Lewis and ABH antigens which is strongly linked to vulnerability of norovirus infection. People known as non-secretors are susceptible to infections caused by *Haemophilus influenzae*, Neisseria meningitides, *Streptococcus pneumoniae* and UTI caused by *E. coli*. ABO blood type is connected to peptic ulcer. The blood type O is highly susceptible to peptic ulcer than other blood type. *Helicobacter pylori* has been implicated to peptic ulcer similarly, *H. pylori* attachment to the human gastric mucosa was mediated by the H type 1 and Le b fucosylated antigens. Soluble glycoproteins of Le b inhibit *H. pylori* binding of *H. pylori*. This justifies the decreased infectivity commonly observed in the blood type A, B, and AB compared to blood group O [14–16].

## **2.2 Coronavirus**

The novel virus, COVID 19 caused by SARS-CoV-2 widely spread around the globe is yet to be fully understood. Factors that influence susceptibility to the disease are age, sex, comorbid chronic disease etc. ABO blood group may influence the susceptibility to COVID. Blood group A have been linked with significant increase risk compared to blood group O due to like virus surface proteins. Blood group O persons can easily recognize these proteins as foreign and by extension confers lower chances to establish the disease [17]. Furthermore, anti-A inhibit binds of glycosylated SARS-CoV S protein expressing cells to angiotensin -converting enzyme 2 on cell membrane thereby truncate the interaction between the virus and its receptors, providing protection. Angiotensin converting enzyme activity is much in blood group B. This explains the possibility non-O blood group have more mortality [18, 19].

#### **2.3 Coagulation**

ABO blood types have been significantly linked to susceptibility to arterial and venous thromboembolism. There is an association between ABO antigens and the structural protein backbone of coagulation factors vWF and factor VIII which affects coagulation. Hypercoagulable plasma potentially causes venous

thromboembolism and is characteristically observed in non- group O individuals due to higher levels of vWF. Von Willebrand factor is a large glycoprotein synthesized by Weibel-palade components in the endothelial cells and alpha granules of platelets. It is the carrier of factor VIII and plays a crucial role in plaletet adhesion and aggregation. Blood group O people have lower levels of vWF due to lack of additional carbohydrate to the terminal sugar. Moreso, plasma vWF is proteolyzed by metalloprotease enzyme ADAMTS13. This is faster in group O than non-O groups vVWF, thereby degrading FVIII levels. Group A, B and AB then have more vWF therefore increases FVIII levels [20].

#### **2.4 Cardiovascular diseases**

Blood group antigens do not cause cardiovascular diseases yet strongly linked to influence susceptibility. The known primary causes of cardiovascular diseases are genetic traits and life style among others. The ATP-binding cassette 2 genes are located at locus 9q34 which plays a significant role in cholesterol regulation. The H antigen has a connection to the structural back bone of coagulation factors (vWF and VIII) glycoproteins. This phenomenon explains greater risk non-O groups have for ischemic heart disease [8, 21].

Preeclampsia is serious condition with leading cause of intrauterine growth restrictions, maternal and foetal morbidity/mortality. Placental protein 13 is galectin that binds to beta-galactoside (N-acetyl-galactosamine, galactose, and fucose) linked to ABO antigens. This protein is observed in a pregnant woman with preeclampsia at early onset [22].

#### **2.5 Malignancy**

There are numerous publications in literature that have reported strong association between red cells antigens and some malignancies. ABH antigens are found on epithelial cells of the GIT, prostate, lungs, breast, uterine cervix, mouth, and bladder and their expression diminishes as malignancy progresses. Blood type antigens play a crucial role in cell signaling, cell recognition and cell adhesion yet these antigens are missing from the red cell membrane glycoprotein/glycolipids of malignant cells. This has been linked to DNA methylation in the promoter region. Blood group A gene in this case may inhibit transcription of the transferase enzyme with resultant loss of A antigen [15, 23]. Factually, malignancy progression results to loss of ABH antigens and potential metastasis of tumor cells. This phenomenon complicates routine red cell typing. Some tumor cells have been observed to mimic blood group A antigen markers giving group A person higher risk of disease progression than non-A blood groups [15]. It is important to note that blood group antigens do not cause malignancy rather susceptibility.

#### **3. Association of other blood types and diseases**

#### **3.1 Haemolytic disease of the foetus and new-born (HDFN)**

Haemolytic disease of the foetus and new born is a major clinical disease associated to Rhesus, Kell, Kidd and Duffy incompatibility between maternal alloantibody (IgG) to foetal antigens with resultant hemolysis of foetal red cells or suppression of the foetal red cell progenitors (commo with Kell system) [24]. Anti-D is the most implicated and severe form of HDFN, yet routine antenatal anti-D prophylaxis has ameliorated it.

*Blood Groups: More than Inheritance of Antigenic Substances - Susceptibility to Some Diseases DOI: http://dx.doi.org/10.5772/intechopen.104593*

## **3.2 Malaria**

In western regions of Africa, it has been reported that individuals with negative Duffy blood type are common and confers protection against malaria caused by *Plasmodium vivax.* The resetting phenomenon commonly seen in parasitized erythrocytes. Forming rosettes is significantly lower in O blood types than blood group A red cells. It was observed that blood type A and B antigens are receptors for resetting on uninfected erythrocytes [25]. Parasitized RBC express resetting which helps us appreciate malaria pathogenesis. In blood group O, plasmodium falciparum invades Rbcs and little rosette is formed yet unstable which binds to uninfected Rbcs to form clusters of cells with resultant narrowing of vascular system. Some gene products such as pfEMP-1 and RIFIN secreted by Rbcs and parasite respectively have been implicated in ABO blood types and susceptibility to malaria among non-group O individuals. This could probably explain the ABO type commonly seen where malaria is prevalent.

## **4. Conclusion**

It is fair to state that blood group antigens are not the primary cause of diseases but are associated to susceptibility to some diseases. Blood group antigens play a role as receptors or ligands to some disease processes. In general, non-O blood types are more susceptible to diseases than O.

## **Author details**

Williams Bitty Azachi1 \* and Kuschak Mathias Dakop2


\*Address all correspondence to: wbitty4@gmail.com

© 2022 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **References**

[1] Fung MK, Grossman BJ, Hillyer CD, Westhoff CM. Technical manual. 18th ed. Bethesda, MD: AABB Press; 2014

[2] Reid ME, Lomas-Francis C, Olsson ML. Blood Group Antigen Facts Book. 3rd ed. Waltham, MA: Academic Press; 2012

[3] Lewis SM, Bain BJ, Bates I, Dacie JV. Dacie and Lewis Practical Haematology. Philadelphia: Churchill Livingstone/ Elsevier; 2012

[4] Cooling L. Blood groups in infection and host susceptibility. Clinical Microbiology Review. 2015;**28**(3): 801-870

[5] Abegaz SB. Human ABO blood groups and their associations with different diseases. BioMed Research International. 2021;**21**:1-9

[6] Boren T, Falk P, Roth KA, Larson G, Normark S. Attachment of helicobacter pylori to human gastric epithelium mediated by blood group antigens. Science. 1993;**262**(5141):1892-1895

[7] Cserti CM, Dzik WH. The ABO blood group system and plasmodium falciparum malaria. Blood. 2007;**110**(7):2250-2258

[8] Anstee DJ. The relationship between blood groups and disease. Blood. 2010;**115**(23):4635-4643

[9] Aird I, Bentall HH, Mehigan JA, Roberts JAF. The blood groups in relation to peptic ulceration and carcinoma of the colon, rectum, breast and bronchus. BMJ. 1954;**2**(4883):315

[10] Clarke CA, Cowan WK, Edwards JW, et al. The relationship of ABO blood groups to duodenal and gastric ulceration. BMJ. 1955;**2**(4940):643

[11] Bjorkholm B, Lundin A, Sillen A, et al. Comparison of genetic divergence and fitness between two subclones of helicobacter pylori. Infection and Immunity. 2001;**69**(12):7832-7838

[12] Cartron JP, Colin Y. Structural and functional diversity of blood group antigens. Transfusion Clinique et Biologique. 2001;**8**(3):163-199

[13] Garratty G. Relationship of Blood Groups to Disease: Do Blood Group Antigens Have a Biological Role? Southern, California: American Red Cross Blood Services; 2005

[14] Ewald R, Sumner S. Blood type biochemistry and human disease, "Wiley interdisciplinary reviews". Systems Biology and Medicine. 2016;**8**(6):517-535

[15] Yamamoto F, Cid E, Yamamoto M, Blancher A. ABO research in the modern era of genomics. Transfusion Medicine Reviews. 2012;**26**(2):103-118

[16] Garratty G. Blood groups and disease: A historical perspective. Transfusion Medicine Reviews. 2000;**14**(4):291-301

[17] Hari. Blood type and COVID-19 risk: O may help, a may hurt. The New England Journal of Medicine. 2020;**11**:325-329

[18] Laguipo ABB. Blood types and COVID-19 risk confirmed. The New England Journal of Medicine. 2020;**136**(18):1888

[19] Chung CM, Wang RY, Chen JW, et al. A genome-wide association study identifies new loci for ACE activity: Potential implications for response to ACE inhibitor. Pharmacogenomics Journal. 2020;**10**:537-544

[20] Franchini M, Capra F, Targher G, Montagnana M, Lippi, g. Relationship between ABO blood group and von Willebrand factor levels: From biology *Blood Groups: More than Inheritance of Antigenic Substances - Susceptibility to Some Diseases DOI: http://dx.doi.org/10.5772/intechopen.104593*

to clinical implications. Thrombosis Journal. 2007;**5**:14

[21] Carpeggiani C, Coceani M, Landi P, Michelassi C, Abbate AL. ABO blood group alleles: A risk factor for coronary artery disease. An angiographic study. Atherosclerosis. 2010;**211**(2):461-466

[22] Than NG, Romero R, Meiri H, et al. P13, maternal ABO blood groups and the risk assessment of pregnancy complications. PLoS One. 2011;**6**(7): e21564

[23] Weisbrod AB, Nilubol N, Weinstein LS, et al. Association of Type-O Blood with neuroendocrine tumors in multiple endocrine neoplasia type 1. The Journal of Clinical Endocrinology & Metabolism. 2013;**98**(1):E109-E114

[24] Vaughan JI, Warwick R, Letsky E. Erythropoietic suppression in foetal anaemia because of Kell alloimmunization. American Journal of Obstetrics and Gynaecology. 1994;**171**(1):247-252

[25] Barragan A, Kremsner PG, Wahlgren M, et al. Blood group a antigen is a coreceptor in *plasmodium falciparum* resetting. Infectious Immunology. 2000;**68**:2971-2975

## **Chapter 6** RH Groups

*Amr J. Halawani*

## **Abstract**

In 1939, a mother gave birth to a stillborn baby and underwent blood transfusion with ABO-matched blood from her husband. This resulted in a hemolytic transfusion reaction (HTR). Levine and Stetson postulated that a novel antigen was present in the baby and father, which was absent in the mother. Therefore, the mother's immune system recognized this antigen and produced antibodies against it. This condition has been known as the hemolytic disease of the newborn for a long period of time. Since the antenatal management of the fetus has been developed, the term has been modified to hemolytic disease of the fetus and newborn (HDFN). This case led to the discovery of the antibody against the first antigen of the RH blood group system, the D antigen. To date, 56 antigens have been recognized within the RH blood group system. The five main antigens are D, C, c, E, and e. As observed in the above-mentioned case, the antibodies against these antigens are implicated in HTR and HDFN.

**Keywords:** RH, anti-D, hemolytic disease of the fetus and newborn, antenatal and postnatal management, anti-D prophylaxis

## **1. Introduction**

The RH blood group system is the most clinically significant blood group system after the ABO blood group system. It is extremely polymorphic, and to date, 56 antigens have been identified and reported by the International Society of Blood Transfusion [1]. The five main antigens of the RH blood group system are D, C, E, c, and e. Other antigens are represented in a combined form, including the ce or f antigen. Some antigens are correlated to specific ethnicities; e.g., the VS antigen is found in the Black population, which is a variant of the e antigen [2].

## **1.1 RH polypeptides**

The antigens of the RH blood group system are encoded by two highly homologous genes, *RHD* and *RHCE*. The cDNA open reading frame of these 2 genes encodes 417 amino acids for each of the RhD and RhCE polypeptides, with a shared sequence identity of 92% (**Figure 1**). The difference between RhD and RhCE polypeptides is 32–35 amino acids, depending on which *RHCE* allele is inherited [*RHce*, *cE*, *Ce*, and *CE*] [3]. The RhD and RhCE polypeptides traverse the membrane lipid bilayer 12 times and form 6 extracellular loops, in which both NH2 and COOH termini are intracellular [4]. In addition, the RhD and RhCE proteins may act as a possible CO2 channel [5].

Normally, the *RHD* gene encodes for the D antigen, whereas the *RHCE* gene encodes for the C, c, E, and e antigens. Eight possible haplotypes have been identified, which vary from one population to another [6]. **Table 1** displays these

#### **Figure 1.**

*The schematic diagram for the RH genes and their proteins. The RHD gene comprises 10 exons (blue boxes). Two RH boxes, upstream and downstream, are flanked by the RHD gene. The RHCE gene has the same number of exons (red boxes), but it is in the opposite orientation. The transmembrane 50A (TMEM50A) gene is flanked by the RHD and RHCE genes, which are indicated by a green arrow. Each protein, RhD and RhCE, traverses the red blood cell membrane 12 times forming 6 extracellular loops. On the RhCE protein, two amino acid substitutions, Ser103Pro and Pro226Ala, are indicated in the second and fourth extracellular loops of the RhCE protein. These gave rise to the antigenic polymorphism of the C/c and E/e antigens.*


#### **Table 1.**

*Frequencies of RH haplotypes in various ethnicities.*

haplotypes along with the frequencies observed in different ethnicities. For instance, *Dce* is the most common haplotype observed in individuals of African origin compared with that observed in individuals from England and southwestern Saudi Arabia [7, 8, 10]. By contrast, the Chinese population lacks the *D* haplotypes [9].

The RhD/RhCE polypeptide is accompanied by two Rh-associated glycoproteins (RhAG) proteins. This association produces a trimer structure as a part of a macrocomplex on the red blood cell membrane proteins [11]. RhAG resembles the RhD and RhCE polypeptides with 36% identity and possesses glycosylation on the first loop. By contrast, the RhD and RhCE polypeptides are not glycosylated [12, 13].

#### **1.2 RHD polymorphisms**

The D+ antigen and D**−** are always denoted as Rh+ and Rh−, respectively. The presence of the D antigen in an individual means that their blood group is D+.

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

However, a person who lacks this antigen is considered D− [14]. Normal D+ individuals have a contact *RHD* gene with 10 contact exons without any mutations or modifications, with RH boxes that flank the *RHD* gene from upstream and downstream [15] (**Figure 1**). Regarding D− individuals, there are various genetic mechanisms underlying this phenotype according to ethnicity. For example, in Caucasians, the entire *RHD* gene in the *dce* haplotype is deleted, resulting in a hybrid box of both upstream and downstream RH boxes [16]. By contrast, Africans possess the pseudogene *RHDΨ*, which has a 37-bp duplication in intron 3 and exon 4, three missense mutations in exon 5, and a nonsense mutation with a premature stop codon in exon 6 [17].

Regarding the Asian population, D− is rare. Nevertheless, different mechanisms have been identified for the D− phenotype in this ethnicity, including the entire deletion of the RHD gene, D-elute (DEL) phenotype, and hybrid genes (such as *RHD-CE(2-9)-D* and *RHD-CE(3-9)-D*) [18–21].

## **1.3 Variants of the D antigen**

## *1.3.1 Weak D*

The weak D phenotype was previously designated as D<sup>u</sup> because it could only be identified by anti-D immunoglobulin (Ig) G in the antiglobulin test and not with anti-D IgM. In contrast to the normal D antigen, the numbers of antigen sites per red blood cell are less and considered quantitative D [22–24]. The intact D antigen (normal D+) has 13,000–24,000 antigen sites per red blood cell. However, the weak D antigen possesses only between 70 and 4000 sites [25].

The weak D antigen comprises all the D epitopes but with weak expression. This phenotype arose from a missense mutation in the intracellular or intramembranous domain of the RhD polypeptides. Thus, a restriction occurs during the RhD polypeptide subunit assembly, leading to a decrease in the density of the RhD polypeptides [26]. In general, individuals with weak D cannot produce anti-D compared with partial D and are treated as D+ individuals [27]. However, in rare scenarios, weak D can produce anti-D. Hence, the term has been modified to weak partial D for such phenotypes. In summary, the term "D variants" was proposed by Daniels to be used for both weak D and partial D to clear the ambiguity [28]. A website called "The Human Rhesus Base" lists all D variant alleles [29, 30].

### *1.3.2 Partial D*

The partial D phenotype was initially classified into six categories (i.e., I–VI) according to the patterns of antibody reactions with D+ red blood cells, which already produced anti-D [31, 32]. The development of monoclonal antibodies paved the way to identify different reaction patterns. The D antigen is now defined as a mosaic or made of pieces of "epitopes." Thirty epitopes have been identified and numbered as ep1–ep9, excluding ep7, followed by the subdivisions of these epitopes (e.g., ep8.3) [33].

This phenotype is characterized by the absence of some epitopes and is considered qualitative D [33]. Such individuals can produce anti-D when undergoing blood transfusion with a "complete" and intact normal D antigen and must be treated as D− phenotype when receiving a blood transfusion. In addition, a D− woman who is pregnant with a complete D+ child who inherits the paternal allele from his father is also at risk of developing hemolytic disease of the fetus and newborn (HDFN) [34]. The gene conversion of the two RH genes leads to the formation of a hybrid gene and the replacement of the *RHD* parts by the corresponding

*RHCE* ones. Furthermore, the presence of a missense mutation in the extracellular domains of the RhD polypeptides could result in the partial D phenotype [35].

#### **1.4 Clinical significance of the RH groups**

The most immunogenic antibody of the RH blood group system is the anti-D, which has been reported to cause severe hemolytic transfusion reaction (HTR). Therefore, typing for the D antigen is extremely crucial, except in a population in which D− is considered rare [36]. Of note, some D− individuals have been reported to produce anti-D antibodies when blood transfusion reaction occurs and D+ blood is transfused [37–39].

Anti-D has also been implicated in severe HDFN, leading to fetal mortality [40]. However, since the start of the use of anti-D prophylaxis, this issue has been decreased dramatically [41]. Other antibodies of the RH blood group system, namely, anti-C, anti-c, anti-E, and anti-e, have been observed to result in severe HTR and HDFN [42]. To date, more than 50 antigens among the RH and different blood group systems have been implicated in HDFN, ranging from mild to severe [43].

## **2. HDFN**

#### **2.1 HDFN pathophysiology**

Maternal alloimmunization may be caused by blood transfusion with an incompatible blood group antigen or during the previous or present pregnancy, in which the fetus or neonate inherits the paternal allele of the blood group antigen that is different from the maternal allele [44]. Fetal red cell leakage via the placenta entering the maternal circulation is known as fetomaternal hemorrhage (FMH). These fetal red cells are recognized by the mother's immune system as foreign bodies and start producing IgM antibodies against these antigens. In the subsequent pregnancy, the maternal IgG antibodies cross the placenta, attach to the fetal red cells, and sensitize them. Consequently, alloimmune destruction can be triggered by splenic macrophages, resulting in anemia with erythroblastosis. The duration of hemolysis varies and causes antenatal or postnatal complications according to the development of the blood group antigen of the fetus [45]. Extramedullary erythropoiesis subsequently occurs to compensate for the destroyed red cells [46].

In the case of mild anemia, an appropriate compensation can be achieved by the liver and spleen. However, in complicated cases, severe anemia leads to hypoxia in multiple organs as a result of difficult delivery that requires sufficient oxygen and nutrients. This subsequently leads to circulatory and liver failure. Liver failure results in a decrease in protein levels and a drop in oncotic pressure in the circulation. Moreover, heart failure increases venous pressure. These two complications lead to ascites and edema, which is recognized as hydrops fetalis that has a high rate of mortality and stillbirth babies [47].

The level of HDFN severity varies from one fetus/infant to another [48]. In most severe cases, the fetus may die in the uterus starting from approximately week 17 of pregnancy [49]. In severe cases presenting with hydrops fetalis, which has high morbidity and mortality, patients could be diagnosed prior to or after the delivery. For the earlier detection of the disease, intrauterine transfusion can be performed in the patients [50]. For moderate HDFN, hyperbilirubinemia can be observed in patients. In such cases, exchange transfusion is needed for the neonate to prevent the accumulation of excess bilirubin in the brain, which may lead to neurological

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

damage and kernicterus. Regarding mild cases, jaundice and hemolysis can be identified in the neonatal period [51].

The hemolysis of the red cells leads to the release of bilirubin into the fetal/neonatal circulation. This released bilirubin has the ability to traverse the placenta, which is then eliminated by the maternal liver. After delivery, the unconjugated bilirubin is processed and excreted by the neonate's liver. The enzyme diphosphate glucuronosyltransferase is released in a lesser amount owing to the immaturity of the neonate's liver [52].

Because of fetal hemoglobin destruction, all newborns can exhibit jaundice. Therefore, newborns with severe hemolysis have an increased level of bilirubin. This excess amount can accumulate in the brain and lead to irreversible central neurological damage and death, a condition known as kernicterus [53].

Anti-D is the major cause of HDFN, and it was a major cause of fetal and neonatal morbidity and mortality before 1970. The incidence of stillbirths and infant deaths dropped after anti-D prophylaxis was developed [54]. This anti-D prophylaxis inhibits the production of the maternal anti-D antibodies after the D-mismatch pregnancies. However, partial D mothers can develop anti-D antibodies when they become pregnant with D+ antigen fetus. In this regard, such mothers need to be treated as D− and should receive anti-D prophylaxis [55].

## **2.2 Factors affecting HDFN immunization and severity**

Many factors affect the severity of RH-HDFN, including antigenic exposure, host factors, Ig class, antibody specificity, and influence of the ABO blood group.

## *2.2.1 Antigenic exposure*

FMH from previous pregnancy can lead to a critical increase in maternal antibody titers, leading to maternal alloimmunization. FMH could occur in approximately 93% of mothers in a small amount (as low as 0.5 mL) [56]. The risk of FMH may be elevated as a result of abdominal trauma or because of interventions, including amniocentesis. Furthermore, the occurrence of FMH may exceed 50% at delivery owing to the entry of fetal red cells into the maternal circulation via placental separation from the uterus [57].

## *2.2.2 Host factors*

With respect to the undefined genetic factors that lead to complications, the capability of antibody production differs in response to antigenic exposure [58]. For example, approximately 85% of D− individuals produce alloanti-D antibodies when transfused with blood containing D+ red cells. However, if anti-D prophylaxis has been administered to D− mothers, the remaining 16% will be at the risk of HDFN after pregnancy with D+ babies [59].

## *2.2.3 Ig class*

Among the Ig classes (IgG, IgM, IgA, IgE, and IgD), only IgG has the ability to cross the placenta. The IgG1 and IgG3 subclasses are more efficient in causing red cell intravascular hemolysis compared with the IgG2 and IgG4 subclasses [60].

## *2.2.4 Antibody specificity*

As previously described in this chapter, among all red cell antibodies, anti-D is the most immunogenic. Moreover, anti-C, anti-c, and anti-E are also potent

antibodies that can cause moderate to severe HDFN. Anti-c and anti-E may lead to severe HDFN, which require management and treatment [61].

#### *2.2.5 Influence of the ABO group*

Of note, the incidence of detecting FMH is decreased when the mother is ABOincompatible with the fetus. The rate of D immunization has been reported to be less in mothers with major ABO incompatibility with the fetus. This may be caused by the clearance of hemolysis that occurs due to the presence of ABO-incompatible D+ fetal red cells in the maternal circulation prior to the identification of fetal D+ red cells by the mother's immune system [57].

#### **2.3 Anti-D prophylaxis**

Anti-D prophylaxis is also known as RH immune globulin. In 1965, the success of the anti-D prophylaxis was reported; the prophylaxis was given to D− mothers after the delivery of the D+ newborn [62]. Anti-D prophylaxis has become the backbone therapy to preclude any clinical significance of RH-HDFN. However, the mode of action regarding how these prophylaxis works remain unclear [63].

Anti-D prophylaxis contains a natural product derived from human plasma that helps in the prevention of sensitization events; therefore, there might be a risk of infectious disease transmission [64]. In general, male blood donors are used and reexposed to a small volume of the D antigen. As a consequence, the anti-D antibody is then developed by the immune system of the donors, followed by plasma collection and processing [65].

Anti-D prophylaxis was only administered to D− mothers postnatally, and in the 1970s, the prophylaxis was modified to include an antenatal dose for the additional prevention of any sensitizing event of RH-HDFN [66].

Different anti-D prophylaxis routines are used in different countries, in which most of them administer the antenatal dose at 34 weeks. The current regimen is by administering two doses to the D− or partial D mothers who are pregnant with D+ babies. The first dose of the anti-D prophylaxis should be administered to D− mothers within 28–34 weeks of pregnancy, which could decrease the antenatal immunization [67, 68]. A total of 92% of mothers were shown to have become sensitized after week 28 [64]. If sensitization events occur, anti-D prophylaxis must be immediately administered within 72 h [69].

The second dose of anti-D prophylaxis must be given within 72 h of delivery to all D− mothers. This dose has reduced the burden of RH-HDFN by approximately 95% in the last 52 years [70]. Indeed, the second dose was initially introduced in the first routine of anti-D prophylaxis [66]. The recommended dosage of anti-D is 300 mg [49]. A larger dose might be given depending on the magnitude of FMH to reduce the risk of sensitization. The dose is typically calculated depending on the magnitude of FMH either using the Kleihauer-Betke test or flow cytometry. Cellular assays might be performed to predict the severity of HDFN, including antibodydependent cellular cytotoxicity assay, monocyte monolayer assay, and chemiluminescence [71].

#### **2.4 Antenatal screening (fetus at risk)**

In general, the recommended routine testing for ABO and D grouping on a maternal blood sample is performed in the first trimester to predict the severity of HDFN. Antibody screening is also performed. Antibody screening must be performed frequently during pregnancy to identify any emerging antibodies.

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

The early investigation of any new antibodies could assist in the monitoring and management of HDFN [72].

Maternal antibody titer testing is performed if IgG antibody screening is positive. Each antibody has a certain titer, which varies as per the antibody. If this titer is below a certain level, further management for HDFN is not necessary [73]. Titer measurement can also be beneficial to distinguish between immune and passive anti-D. Nevertheless, the severity of HDFN cannot be reliably determined [74]. If the maternal antibody level is higher than the critical titer, paternal *RHD* zygosity testing may be required.

## *2.4.1 Predicting D phenotype from DNA*

## *2.4.1.1 Paternal RHD zygosity testing*

The management of all immunized mothers is possible via paternal *RHD* zygosity testing, which detects the copy number of the father's gene. This identifies the father's zygosity status and paternity. Nowadays, various techniques have been used for *RHD* zygosity testing. These include real-time polymerase chain reaction (PCR), digital PCR, and mass spectrometry [75–78].

When fathers are homozygous for the deletion of the entire *RHD* gene, i.e., D−, there is no risk of HDFN within the current or subsequent pregnancies. By contrast, when fathers are D+ carrying both the alleles of positive *RHD* gene (homozygous, e.g., *DCe*/*DCe*), then the fetus will definitely be D+. The fetus has a 50% possibility of being D+ in case the father's genotype is hemizygous (e.g., *DCe*/*dce*). Therefore, further analysis using noninvasive prenatal testing (NIPT) is required [77].

Some methods assess the hybrid Rhesus box expressed in D− individuals, which results from the entire *RHD* gene deletion in Caucasians. It mainly targets the *cde* haplotype; therefore, such a method cannot be applied to the samples of African ethnicity [79]. A total of 59% of Nigerians [8] possess the *Dce* haplotype [see **Table 1**].

## *2.4.1.2 Fetal genotyping*

Amniocentesis is a risky and invasive procedure for obtaining fetal DNA. It can increase the risk of alloimmunization and miscarriage. Instead, NIPT is nowadays performed for fetal genotyping. The approach is now followed globally and is performed by detecting cell-free fetal DNA extracted from the maternal plasma [80, 81].

Fetal genotyping is a noninvasive technique based on cell-free fetal DNA circulating in the maternal plasma, which is derived from a maternal blood sample. This approach assists in identifying different blood group alleles and predicting the corresponding antigens, including D, C, c, E, and e, in addition to KEL1 antigens [82, 83].

If the mother is D− and the fetus is D+, anti-D prophylaxis may be administered. By contrast, if the fetus is D−,the D− mother does not need to receive the prophylaxis. Therefore, this avoids unnecessary injections in the mother; moreover, it appears to be cost-effective to save treatment for the mothers in actual need [84]. Thus, the approach of fetal genotyping could be cost-effective [85].

## **2.5 Antenatal management**

## *2.5.1 Ultrasonography*

Amniocentesis has been replaced by the middle cerebral artery (MCA) Doppler ultrasonography for predicting the severity of fetal anemia. At present, this method is routinely used by obstetricians for investigating fetal anemia by observing an increase in the velocity of blood flow in MCA in anemic fetuses compared with normal ones [86, 87]. In the case of severe anemia, fetal blood sampling (FBS) can be performed along with cordocentesis. Under ultrasound guidance, FBS is normally performed with a needle to obtain fetal blood; this procedure is considered an invasive procedure [88].

### *2.5.2 Intrauterine transfusion*

For severely HDFN-affected fetuses, intrauterine transfusion provides blood to the fetus via the umbilical vein under ultrasound guidance. Fetuses must be administered O**–** blood (unless fetal ABO type is known), which is also KEL-1 negative, leuko depleted, plasma depleted, hemoglobin S negative, cytomegalovirus seronegative, and ≤5 days old [89]. Furthermore, citrate phosphate dextrose is the anticoagulant used in these blood units to prevent problems arising from using different anticoagulants. Gamma irradiation is performed to eliminate any residual leukocytes that may lead to graft-versus-host disease [89].

Fetal hemoglobin and hematocrit level measurement along with crossmatching is performed to ensure that safe compatible blood is transfused to the fetus. All blood units are normally prewarmed to 37°C before being transfused. Interestingly, intrauterine transfusion has been reported to reduce prenatal death and stillbirth resulting from HDFN by 75–90% [90, 91].

### **2.6 Postnatal screening (newborn/infant at risk)**

The serological investigation is performed after a sample is withdrawn from the cord blood at birth. This sample is used to detect HDFN, which may help arrange for probable transfusion.

#### *2.6.1 ABO grouping*

The forward ABO blood group typing may be observed with weak reactions with anti-A and anti-B antisera in infants compared with adults and older children [57]. This is because the ABO antigens of newborn infants are not entirely developed and may take 5**–**10 years to reach the adult levels [92]. Furthermore, reverse ABO grouping is not feasible because infants do not produce ABO antibodies at that age.

#### *2.6.2 D typing*

In rare cases, an infant's red cells strongly bind to the maternal anti-D antibody, resulting in a false-negative type of D or what is known as blocked RH [93]. Anti-D can be identified from the eluant of these red cells, and typing these eluted red blood cells should be observed for any reaction with the anti-D antibody.

#### *2.6.3 Direct antiglobulin test (DAT)*

DAT is very crucial in diagnosing HDFN. A positive DAT demonstrates a sensitization reaction in which an infant's red cells are coated by maternal IgG antibodies. No correlation has been observed between the severity of HDFN and the reaction strength. Moreover, other laboratory or clinical manifestations for hemolysis can also lead to positive DAT. This could be owing to the mother receiving the anti-D prophylaxis [57].

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

## *2.6.4 Elution*

Performing the elution test in the case of positive DAT is not essential as a routine procedure. As previously mentioned, eluant is the solution for blocked RH [57, 93].

## **2.7 Infant management**

### *2.7.1 Phototherapy*

This procedure is used to treat anemic newborns with mild to moderate HDFN and who have elevated levels of bilirubin. Phototherapy is performed using a blue**-**green light, with wavelength ranging from 460 to 490 nm [94]. Natural direct sunlight has been reported to have the same wavelength and can be beneficial to reduce jaundice, although it is not recommended because it may increase the risk of sunburn [95, 96].

Bilirubin is a lipophilic molecule that absorbs light and is metabolized into two isomers, which are less lipophilic (in other words, water-soluble) and less toxic to the brain. These can then be excreted via the urine without the requirement of enzymatic glucuronidation [97]. Overall, phototherapy is an effective procedure and can adequately conjugate bilirubin. Furthermore, it may assist reduce the requirement of blood transfusion [57].

#### *2.7.2 Exchange transfusion*

Exchange transfusion may be performed in newborns who demonstrate severe anemia with hyperbilirubinemia or heart failure. During pregnancy, the fetal liver is unable to metabolize the unconjugated bilirubin. Therefore, this unconjugated bilirubin, made by the fetus, is crossed the placenta and metabolized by the maternal disposal system. After delivery, this system is no longer used as well as the infant's liver is immature and cannot metabolize the unconjugated bilirubin efficiently. Therefore, the high level of bilirubin in the newborn may lead to kernicterus, which is the accumulation of bilirubin in the brain.

The assessment of hemoglobin and bilirubin levels is essential to determine the requirement of exchange transfusion in neonates. This is performed for removing bilirubin and maternal antibodies from neonates [98, 99]. Exchange transfusion is indicated at a critical level of bilirubin (i.e., ≥100 μmol/L), depending on the neonatal age.

Exchange transfusion is the replacement of neonatal blood by whole blood or equivalent, with the concurrent removal of bilirubin and maternal antibodies. However, this procedure is labor-intensive and time-consuming. Therefore, its use has become rare owing to the use of anti-D prophylaxis and phototherapy [50].

#### *2.7.3 Red cell transfusion*

Infants may receive red cell transfusion immediately after delivery for several weeks to treat severe anemia. The same criteria that are used for intrauterine transfusion and exchange transfusion must be applied for blood transfusion. Newborns must be closely monitored for any clinical signs of ongoing anemia, particularly if the infant is malnourished or sleeps heavily [98].

## **3. Conclusion**

In summary, RH is the most highly variable blood group system. The antigens of this system have many variants that include the D variants such as weak D and partial D. The antibodies of this system can cause HTR due to incompatibility during a blood transfusion. Furthermore, their antibodies can lead to fetal red cell sensitization and destruction, causing moderate to severe HDFN. Nowadays, owing to the development of the latest technologies, the antenatal management of fetuses has become feasible along with postnatal management. Anti-D prophylaxis can now be administered antenatally at gestational weeks 28–34. At-risk pregnancies can also be monitored noninvasively using MCA ultrasonography and fetal genotyping.

## **Conflict of interest**

The author declares no conflict of interest.

## **Author details**

Amr J. Halawani Faculty of Applied Medical Sciences, Department of Laboratory Medicine, Umm Al-Qura University, Makkah, Saudi Arabia

\*Address all correspondence to: ajjhalawani@uqu.edu.sa

© 2022 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **References**

[1] International Society of Blood Transfusion. Red cell immunogenetics and blood group terminology [Internet]. 2021. Available from: https://www. isbtweb.org/working-parties/red-cellimmunogenetics-and-blood-groupterminology [Accessed: September 06, 2021]

[2] Chou ST, Westhoff CM. The Rh and RhAG blood group systems. Immunohematology. 2010;**26**:178-186. Available from: https://pubmed.ncbi. nlm.nih.gov/22356455/

[3] Mouro I, Colin Y, Chérif-Zahar B, Cartron JP, Le Van Kim C. Molecular genetic basis of the human rhesus blood group system. Nature Genetics. 1993;**5**:62-65. DOI: 10.1038/ng0993-62

[4] Chérif-Zahar B, Bloy C, Le Van Kim C, Blanchard D, Bailly P, Hermand P, et al. Molecular cloning and protein structure of a human blood group Rh polypeptide. Proceedings of the National Academy of Sciences of the United States of America. 1990;**87**:6243- 6247. DOI: 10.1073/pnas.87.16.6243

[5] Soupene E, King N, Feild E, Liu P, Niyogi KK, Huang CH, et al. Rhesus expression in a green alga is regulated by CO2. Proceedings of the National Academy of Sciences of the United States of America. 2002;**99**:7769-7773. DOI: 10.1073/pnas.112225599

[6] Daniels G. Rh and RHAG blood group systems. In: Human Blood Groups. 3rd ed. UK: Wiley-Blackwell; 2013. DOI: 10.1002/9781118493595.ch5

[7] Race RR, Mourant AE, Lawler SD, Sanger R. The Rh chromosome frequencies in England. Blood. 1948;**3**:689-695. DOI: 10.1182/blood. V3.6.689.689

[8] Mourant AE, Domaniewska-Sobczak K, Kopec AC. The Distribution of the Human Blood Groups and Other Polymorphisms. 2nd ed. London: Oxford University Press; 1976. Available from: http://lib.ugent.be/catalog/ rug01:000041013

[9] Mackay J, Wang JC, Wong KS. The incidence of blood groups in 4648 southern Chinese from Toishan district and its vicinity. Journal of Hong Kong Medical Technology Association. 1969;**1**:11-14

[10] Halawani AJ, Arjan AH. ABO, RH, and KEL1 antigens, phenotypes and haplotypes in southwestern Saudi Arabia. Clinical Laboratory. 2021;**67**: 344-348. DOI: 10.7754/Clin.Lab. 2020.200633

[11] Conroy MJ, Bullough PA, Merrick M, Avent ND. Modelling the human rhesus proteins: Implications for structure and function. British Journal of Haematology. 2005;**131**:543-551. DOI: 10.1111/j.1365-2141.2005.05786.x

[12] Eyers SA, Ridgwell K, Mawby WJ, Tanner MJ. Topology and organization of human Rh (rhesus) blood grouprelated polypeptides. The Journal of Biological Chemistry. 1994;**269**:6417- 6423. DOI: 10.1016/S0021-9258 (17)37388-X

[13] Marini AM, Urrestarazu A, Beauwens R, André B. The Rh (rhesus) blood group polypeptides are related to NH4 + transporters. Trends in Biochemical Sciences. 1997;**22**:460-461. DOI: 10.1016/s0968-0004(97)01132-8

[14] Colin Y, Chérif-Zahar B, Le Van Kim C, Raynal V, Van Huffel V, Cartron JP. Genetic basis of the RhDpositive and RhD-negative blood group polymorphism as determined by Southern analysis. Blood. 1991;**78**:2747- 2752. DOI: 10.1182/blood.V78.10. 2747.2747

[15] Chérif-Zahar B, Le Van Kim C, Rouillac C, Raynal V, Cartron JP, Colin Y. Organization of the gene (RHCE) encoding the human blood group RhCcEe antigens and characterization of the promoter region. Genomics. 1994;**19**:68-74. DOI: 10.1006/ geno.1994.1014

[16] Wagner FF, Flegel WA. RHD gene deletion occurred in the rhesus box. Blood. 2000;**95**:3662-3668. DOI: 10.1182/blood.V95.12.3662

[17] Singleton BK, Green CA, Avent ND, Martin PG, Smart E, Daka A, et al. The presence of an RHD pseudogene containing a 37 base pair duplication and a nonsense mutation in Africans with the Rh D-negative blood group phenotype. Blood. 2000;**95**:12-18. DOI: 10.1182/blood.V95.1.12

[18] Shao CP, Maas JH, Su YQ, Köhler M, Legler TJ. Molecular background of Rh D-positive, D-negative, D(el) and weak D phenotypes in Chinese. Vox Sanguinis. 2002;**83**:156-161. DOI: 10.1046/j.1423-0410.2002.00192.x

[19] Sun CF, Chou CS, Lai NC, Wang WT. RHD gene polymorphisms among RhD-negative Chinese in Taiwan. Vox Sanguinis. 1998;**75**:52-57. DOI: 10.1159/000030958

[20] Peng CT, Shih MC, Liu TC, Lin IL, Jaung SJ, Chang JG. Molecular basis for the RhD negative phenotype in Chinese. International Journal of Molecular Medicine. 2003;**11**:515-521. DOI: 10.3892/ijmm.11.4.515

[21] Xu Q, Grootkerk-Tax MG, Maaskant-van Wijk PA, Van Der Schoot CE. Systemic analysis and zygosity determination of the RHD gene in a D-negative Chinese Han population reveals a novel D-negative RHD gene. Vox Sanguinis. 2005;**88**:35-40. DOI: 10.1111/j.1423-0410.2005.00584.x

[22] Bush M, Sabo B, Stroup M, Masouredis SP. Red cell D antigen sites and titration scores in a family with weak and normal Du phenotypes inherited from a homozygous Du mother. Transfusion. 1974;**14**:433-439. DOI: 10.1111/j.1537-2995.1974.tb04557.x

[23] Cunningham NA, Zola AP, Hui HL, Taylor LM, Green FA. Binding characteristics of anti-Rh0 (D) antibodies to Rh0 (D)-positive and Du red cells. Blood. 1985;**66**:765-768. DOI: 10.1182/blood.V66.4.765.765

[24] Szymanski IO, Araszkiewicz P. Quantitative studies on the D antigen of red cells with the Du phenotype. Transfusion. 1989;**29**:103-105. DOI: 10.1046/j.1537-2995.1989.29289146825.x

[25] Wagner FF, Frohmajer A, Ladewig B, Eicher NI, Lonicer CB, Müller TH, et al. Weak D alleles express distinct phenotypes. Blood. 2000;**95**:2699-2708. DOI: 10.1182/blood. V95.8.2699

[26] Wagner FF, Gassner C, Müller TH, Schönitzer D, Schunter F, Flegel WA. Molecular basis of weak D phenotypes. Presented at the 25th Congress of the International Society of Blood Transfusion held in Oslo on June 29, 1998 and published in abstract form in Vox Sang 74:55, 1998[suppl]. Blood. 1999;**93**(1):385-393. DOI: 10.1182/blood. V93.1.385

[27] Sandler SG, Chen LN, Flegel WA. Serological weak D phenotypes: A review and guidance for interpreting the RhD blood type using the RHD genotype. British Journal of Haematology. 2017;**179**:10-19. DOI: 10.1111/bjh.14757

[28] Daniels G. Variants of RhD--current testing and clinical consequences. British Journal of Haematology. 2013;**161**:461- 470. DOI: 10.1111/bjh.12275

[29] The human RhesusBase, version 2.5. Update. 2020. Available from: http:// www.rhesusbase.info/

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

[30] Wagner FF, Flegel WA. The rhesus site. Transfusion Medicine and Hemotherapy. 2014;**41**:357-363. DOI: 10.1159/000366176

[31] Tippett P, Sanger R. Observations on subdivisions of the Rh antigen D. Vox Sanguinis. 1962;**7**:9-13. DOI: 10.1111/ j.1423-0410.1962.tb03223.x

[32] Tippett P. Further observations on subdivisions of the Rh antigen D. 1977. Available from: http://pascal-francis. inist.fr/vibad/index.php?action=getRec ordDetail&idt=PASCAL7850148289

[33] Scott ML, Voak D, Jones JW, Avent ND, Liu W, Hughes-Jones N, et al. A structural model for 30 Rh D epitopes based on serological and DNA sequence data from partial D phenotypes. Transfusion Clinique et Biologique. 1996;**3**:391-396. DOI: 10.1016/ S1246-7820(96)80051-6

[34] Argall CI, Ball JM, Trentelman E. Presence of anti-D antibody in the serum of a Du patient. The Journal of Laboratory and Clinical Medicine. 1953;**41**:895-898. DOI: 10.5555/ uri:pii:0022214353900730

[35] Flegel WA. Molecular genetics of RH and its clinical application. Transfusion Clinique et Biologique. 2006;**13**:4-12. DOI: 10.1016/j. tracli.2006.02.011

[36] Lin M. Taiwan experience suggests that RhD typing for blood transfusion is unnecessary in Southeast Asian populations. Transfusion. 2006;**46**:95- 98. DOI: 10.1111/j.1537-2995.2006. 00680.x

[37] Frohn C, Dümbgen L, Brand JM, Görg S, Luhm J, Kirchner H. Probability of anti-D development in D− patients receiving D+ RBCs. Transfusion. 2003;**43**:893-898. DOI: 10.1046/j. 1537-2995.2003.00394.x

[38] Yazer MH, Triulzi DJ. Detection of anti-D in D−recipients transfused with D+ red blood cells. Transfusion. 2007;**47**:2197-2201. DOI: 10.1111/j. 1537-2995.2007.01446.x

[39] Gonzalez-Porras JR, Graciani IF, Perez-Simon JA, Martin-Sanchez J, Encinas C, Conde MP, et al. Prospective evaluation of a transfusion policy of D+ red blood cells into D− patients. Transfusion. 2008;**48**:1318-1324. DOI: 10.1111/j.1537-2995.2008.01700.x

[40] Levine P, Stetson RE. An unusual case of intra-group agglutination. Journal of the American Medical Association. 1939;**113**:126-127. DOI: 10.1001/jama.1939.72800270002007a

[41] Freda VJ, Gorman JG, Pollack W. Rh factor: Prevention of isoimmunization and clinical trial on mothers. Science. 1966;**151**:828-830. DOI: 10.1126/ science.151.3712.828

[42] Koelewijn JM, Vrijkotte TG, Van Der Schoot CE, Bonsel GJ, De Haas M. Effect of screening for red cell antibodies, other than anti-D, to detect hemolytic disease of the fetus and newborn: A population study in the Netherlands. Transfusion. 2008;**48**:941-952. DOI: 10.1111/j.1537-2995.2007.01625.x

[43] Moise KJ. Hemolytic disease of the fetus and newborn. Clinical Advances in Hematology & Oncology. 2013; **11**:664-666

[44] De Haas M, Thurik FF, Koelewijn JM, Van der Schoot CE. Haemolytic disease of the fetus and newborn. Vox Sanguinis. 2015;**109**:99- 113. DOI: 10.1111/vox.12265

[45] Lopriore E, Rath ME, Liley H, Smits-Wintjens VE. Improving the management and outcome in haemolytic disease of the foetus and newborn. Blood Transfusion. 2013;**11**:484

[46] Urbaniak SJ, Greiss MA. RhD haemolytic disease of the fetus and the newborn. Blood Reviews. 2000;**14**:44- 61. DOI: 10.1054/blre.1999.0123

[47] Smits-Wintjens VE, Walther FJ, Lopriore E. Rhesus haemolytic disease of the newborn: Postnatal management, associated morbidity and long-term outcome. Seminars in Fetal and Neonatal Medicine. 2008;**13**:265-271. DOI: 10.1016/j.siny.2008.02.005

[48] Dean L. Chapter 4, Hemolytic disease of the newborn. In: Blood Groups and Red Cell Antigens [Internet]. Bethesda, US: National Center for Biotechnology Information; 2005. Available from: https://www.ncbi. nlm.nih.gov/books/NBK2266/

[49] Klein HG, Anstee DJ. Chapter 12—Haemolytic disease of the foetus and newborn. In: Mollison's Blood Transfusion in Clinical Medicine. 12th ed. UK: John Wiley & Sons, Wiley-Blackwell; 2014

[50] Jackson ME, Baker JM. Hemolytic disease of the fetus and newborn: Historical and current state. Clinics in Laboratory Medicine. 2021;**41**:133-151. DOI: 10.1016/j.cll.2020.10.009

[51] Akkök ÇA, Seghatchian J. Pediatric red cell and platelet transfusions. Transfusion and Apheresis Science. 2018;**57**:358-362. DOI: 10.1016/j. transci.2018.05.019

[52] Mitchell S, James A. Severe late anemia of hemolytic disease of the newborn. Paediatrics & Child Health. 1999;**4**:201-203. DOI: 10.1093/ pch/4.3.201

[53] Leung AK, Sauve RS. Breastfeeding and breast milk jaundice. Journal of the Royal Society of Health. 1989;**109**:213- 217. DOI: 10.1177/146642408910900615

[54] Tovey LA. Oliver memorial lecture. Towards the conquest of Rh haemolytic disease: Britain's contribution and the role of serendipity. Transfusion

Medicine. 1992;**2**:99-109. DOI: 10.1111/ j.1365-3148.1992.tb00142.x

[55] Lukacevic Krstic JL, Dajak S, Bingulac-Popovic J, Dogic V, Mratinovic-Mikulandra J. Anti-D antibodies in pregnant D variant antigen carriers initially typed as RhD+. Transfusion Medicine and Hemotherapy. 2016;**43**:419-424. DOI: 10.1159/000446816

[56] Sebring ES, Polesky HF. Fetomaternal hemorrhage: Incidence, risk factors, time of occurrence, and clinical effects. Transfusion. 1990;**30**:344-357. DOI: 10.1046/j. 1537-2995.1990.30490273444.x

[57] Harmening DM. Hemolytic disease of the fetus and newborn (HDFN). In: Modern Blood Banking & Transfusion Practices. 7th ed; Philadelphia: FA. Davis Company; 2018

[58] Higgins JM, Sloan SR. Stochastic modeling of human RBC alloimmunization: Evidence for a distinct population of immunologic responders. Blood. 2008;**112**:2546-2553. DOI: 10.1182/blood-2008-03-146415

[59] Ayache S, Herman JH. Prevention of D sensitization after mismatched transfusion of blood components: Toward optimal use of RhIG. Transfusion. 2008;**48**:1990-1999. DOI: 10.1111/j.1537-2995.2008.01800.x

[60] Singh A, Solanki A, Chaudhary R. Demonstration of IgG subclass (IgG1 and IgG3) in patients with positive direct antiglobulin tests. Immunohematology. 2014;**30**:24-27

[61] Babinszki A, Berkowitz RL. Haemolytic disease of the newborn caused by anti-c, anti-E and anti-Fya antibodies: Report of five cases. Prenatal Diagnosis. 1999;**19**:533-536. DOI: 10.1002/(SICI)1097-0223 (199906)19:6<533::AID-PD570>3. 0.CO;2-5

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

[62] Freda VJ, Gorman JG, Pollack W. Successful prevention of experimental Rh sensitization in man with an anti-Rh gamma 2-globulin antibody preparation: A preliminary report. Transfusion. 1964;**4**:26-32. DOI: 10.1111/j.1537- 2995.1964.tb02824.x

[63] Kumpel BM, Elson CJ. Mechanism of anti-D-mediated immune suppression–a paradox awaiting resolution? Trends in Immunology. 2001;**22**:26-31. DOI: 10.1016/S1471- 4906(00)01801-9

[64] Bowman JM. RhD hemolytic disease of the newborn. The New England Journal of Medicine. 1998;**339**:1775- 1777. DOI: 10.1056/NEJM1998121 03392410

[65] Kumpel BM. Monoclonal anti-D for prophylaxis of RhD haemolytic disease of the newborn. Transfusion Clinique et Biologique. 1997;**4**:351-356. DOI: 10.1016/s1246-7820(97)80040-7

[66] Liumbruno GM, D'Alessandro A, Rea F, Piccinini V, Catalano L, Calizzani G, et al. The role of antenatal immunoprophylaxis in the prevention of maternal-foetal anti-Rh (D) alloimmunisation. Blood Transfusion. 2010;**8**:8-16. DOI: 10.2450/2009.0108-09

[67] Jones ML, Wray J, Wight J, Chilcott J, Forman K, Tappenden P, et al. A review of the clinical effectiveness of routine antenatal anti-D prophylaxis for rhesus-negative women who are pregnant. BJOG : An International Journal of Obstetrics and Gynaecology. 2004;**111**:892-902. DOI: 10.1111/j. 1471-0528.2004.00243.x

[68] Kumar S, Regan F. Management of pregnancies with RhD alloimmunisation. BMJ. 2005;**330**:1255-1258. DOI: 10.1136/bmj.330.7502.1255

[69] Crowther CA, Middleton P. Anti-D administration after childbirth for preventing rhesus alloimmunisation.

Cochrane Database of Systematic Reviews. 2000;**2**:CD000021. DOI: 10.1002/14651858.CD000021

[70] Weatherall DJ. Cyril Clarke and the prevention of rhesus haemolytic disease of the newborn. British Journal of Haematology. 2012;**157**:41-46. DOI: 10.1111/j.1365-2141.2012.09031.x

[71] Hadley AG. Laboratory assays for predicting the severity of haemolytic disease of the fetus and newborn. Transplant Immunology. 2002;**10**:191- 198. DOI: 10.1016/S0966-3274(02) 00065-5

[72] Fung MK, Grossman BJ, Hillyer CD, Westhoff CM, editors. Technical Manual. 18th ed. Bethesda: American Association of Blood Banks Press; 2014

[73] Bennardello F, Coluzzi S, Curciarello G, Todros T, Villa S, Italian Society of Transfusion Medicine and Immunohaematology (SIMTI) and Italian Society of Gynaecology and Obstetrics (SIGO) Working Group. Recommendations for the prevention and treatment of haemolytic disease of the foetus and newborn. Blood Transfusion. 2015;**13**:109-134. DOI: 10.2450/2014.0119-14

[74] Gottvall T, Hllden JO. Concentration of anti-D antibodies in Rh (D) alloimmunized pregnant women, as a predictor of anemia and/or hyperbilirubinemia in their newborn infants. Acta Obstetricia et Gynecologica Scandinavica. 1997;**76**:733-738. DOI: 10.3109/ 00016349709024338

[75] Chiu RW, Murphy MF, Fidler C, Zee BC, Wainscoat JS, Lo YM. Determination of RhD zygosity: Comparison of a double amplification refractory mutation system approach and a multiplex real-time quantitative PCR approach. Clinical Chemistry. 2001;**47**:667-672. DOI: 10.1093/ clinchem/47.4.667

[76] Krog GR, Clausen FB, Dziegiel MH. Quantitation of RHD by real-time polymerase chain reaction for determination of RHD zygosity and RHD mosaicism/chimerism: An evaluation of four quantitative methods. Transfusion. 2007;**47**:715-722. DOI: 10.1111/j.1537-2995.2007.01175.x

[77] Sillence KA, Halawani AJ, Tounsi WA, Clarke KA, Kiernan M, Madgett TE, et al. Rapid RHD zygosity determination using digital PCR. Clinical Chemistry. 2017;**63**:1388-1397. DOI: 10.1373/clinchem.2016.268698

[78] Gassner C, Meyer S, Frey BM, Vollmert C. Matrix-assisted laser desorption/ionisation, time-of-flight mass spectrometry–based blood group genotyping—the alternative approach. Transfusion Medicine Reviews. 2013;**27**:2-9. DOI: 10.1016/j. tmrv.2012.10.001

[79] Grootkerk-Tax MG, Maaskant-van Wijk PA, Van Drunen J, Van Der Schoot CE. The highly variable RH locus in nonwhite persons hampers RHD zygosity determination but yields more insight into RH-related evolutionary events. Transfusion. 2005;**45**:327-337. DOI: 10.1111/j.1537-2995.2005.04199.x

[80] Norwitz ER, Levy B. Noninvasive prenatal testing: The future is now. Reviews in Obstetrics and Gynecology. 2013;**6**:48-62

[81] Chandrasekharan S, Minear MA, Hung A, Allyse MA. Noninvasive prenatal testing goes global. Science Translational Medicine. 2014;**6**(6):231fs15. DOI: 10.1126/ scitranslmed.3008704

[82] Finning K, Martin P, Summers J, Daniels G. Fetal genotyping for the K (Kell) and Rh C, c, and E blood groups on cell-free fetal DNA in maternal plasma. Transfusion. 2007;**47**:2126- 2133. DOI: 10.1111/j.1537-2995. 2007.01437.x

[83] Finning KM, Martin PG, Soothill PW, Avent ND. Prediction of fetal D status from maternal plasma: Introduction of a new noninvasive fetal RHD genotyping service. Transfusion. 2002;**42**:1079-1085. DOI: 10.1046/j. 1537-2995.2002.00165.x

[84] Tiblad E, Taune Wikman A, Ajne G, Blanck A, Jansson Y, Karlsson A, et al. Targeted routine antenatal anti-D prophylaxis in the prevention of RhD immunisation-outcome of a new antenatal screening and prevention program. PLoS One. 2013;**8**:e70984. DOI: 10.1371/journal.pone.0070984

[85] Teitelbaum L, Metcalfe A, Clarke G, Parboosingh JS, Wilson RD, Johnson JM. Costs and benefits of non-invasive fetal RhD determination. Ultrasound in Obstetrics & Gynecology. 2015;**45**:84- 88. DOI: 10.1002/uog.14723

[86] El Shourbagy S, Elsakhawy M. Prediction of fetal anemia by middle cerebral artery Doppler. Middle East Fertility Society Journal. 2012;**17**:275- 282. DOI: 10.1016/j.mefs.2012.09.003

[87] Mari G, Deter RL, Carpenter RL, Rahman F, Zimmerman R, Moise KJ Jr, et al. Noninvasive diagnosis by Doppler ultrasonography of fetal anemia due to maternal red-cell alloimmunization. The New England Journal of Medicine. 2000;**342**:9-14. DOI: 10.1056/ NEJM200001063420102

[88] Avent ND. Haemolytic disease of the fetus and newborn. In: Transfusion and Transplantation Science. UK: Oxford University Press; 2018. Available from: https://books.google.com.sa/ books?id=A\_okvgAACAAJ

[89] Oepkes D, van Scheltema PA. Intrauterine fetal transfusions in the management of fetal anemia and fetal thrombocytopenia. Seminars in Fetal & Neonatal Medicine. 2007;**12**:432-438. DOI: 10.1016/j.siny.2007.06.007

#### *RH Groups DOI: http://dx.doi.org/10.5772/intechopen.102421*

[90] Rodeck CH, Holman CA, Karnicki J, Kemp JR, Whitmore DN, Austin MA. Direct intravascular fetal blood transfusion by fetoscopy in severe rhesus isoimmunisation. Lancet. 1981;**317**:625-627. DOI: 10.1016/ S0140-6736(81)91549-X

[91] Kanhai HH, Bennebroek Gravenhorst JB, Van Kamp IL, Meerman RH, Brand A, Dohmen-Feld MW, et al. Management of severe hemolytic disease with ultrasound-guided intravascular fetal transfusions. Vox Sanguinis. 1990;**59**:180-184. DOI: 10.1111/j.1423- 0410.1990.tb00855.x

[92] Auf der Maur CA, Hodel M, Nydegger UE. Rieben RAge dependency of ABO histo-blood group antibodies: Reexamination of an old dogma. Transfusion. 1993;**33**:915-918. DOI: 10.1046/j.1537-2995.1993. 331194082382.x

[93] Sulochana PV, Rajesh A, Mathai J, Sathyabhama S. Blocked D phenomenon, a rare condition with Rh D haemolytic disease of newborn–a case report. International Journal of Laboratory Hematology. 2008;**30**:244- 247. DOI: 10.1111/j.1751-553X.2007. 00943.x

[94] Ebbesen F, Hansen TWR, Maisels MJ. Update on phototherapy in jaundiced neonates. Current Pediatric Reviews. 2017;**13**:176-180. DOI: 10.2174/ 1573396313666170718150056

[95] Salih FM. Can sunlight replace phototherapy units in the treatment of neonatal jaundice? An in vitro study. Photodermatology, Photoimmunology & Photomedicine. 2001;**17**:272-277. DOI: 10.1034/j.1600-0781.2001.170605.x

[96] Slusher TM, Olusanya BO, Vreman HJ, Wong RJ, Brearley AM, Vaucher YE, et al. Treatment of neonatal jaundice with filtered sunlight in Nigerian neonates: Study protocol of a

non-inferiority, randomized controlled trial. Trials. 2013;**14**:446. DOI: 10.1186/1745-6215-14-446

[97] Bhutani VK, Stark AR, Lazzeroni LC, Poland R, Gourley GR, Kazmierczak S, et al. Predischarge screening for severe neonatal hyperbilirubinemia identifies infants who need phototherapy. The Journal of Pediatrics. 2013;**162**:477-482.e1. DOI: 10.1016/j.jpeds.2012.08.022

[98] Rath ME, Smits-Wintjens VE, Walther FJ, Lopriore E. Hematological morbidity and management in neonates with hemolytic disease due to red cell alloimmunization. Early Human Development. 2011;**87**:583-588. DOI: 10.1016/j.earlhumdev.2011.07.010

[99] American Academy of Pediatrics Subcommittee on Hyperbilirubinemia. Management of hyperbilirubinemia in the newborn infant 35 or more weeks of gestation. Pediatrics. 2004;**114**:297-316. DOI: 10.1542/peds.114.1.297

## **Chapter 7**
