Serological Characteristics of Subtype A Caused by New A Allele Mutation and a Family Survey / 中国实验血液学杂志

OBJECTIVE@#In this study, the results of forward and reverse blood typing of a male patient diagnosed as bronchiectasis were inconsistent, which were type O and type A respectively. Multiple experiments including genotyping and sequencing and family investigation were carried out to determine the subtype of ABO blood group and explore the serological characteristics of this subtype.@*METHODS@#Standard serological techniques were used to conduct forward and reverse typing, reverse blood typing enhancement test, H antigen identification, absorption-elution test, salivary blood group substances test, and PCR-SSP method for ABO genotyping and exon 6 and 7 sequencing.@*RESULTS@#The proband's blood group was type O by forward typing, but antigen A could be detected by absorption-elution test, anti-A1 could be detected by reverse blood typing enhancement test, it was found that there was substance H but no substance A in saliva, and the serological characteristics were consistent with Ael subtype. Gene sequencing analysis showed that there was a c.625T>G base substitution on the basis of A102, which had never been reported before. Family survey showed that c.625T>G base substitution appeared in three generations of the family.@*CONCLUSION@#In this study, a new subtype A with Ael serological characteristics caused by c.625T>G mutation was identified. c.625T>G base substitution results in the weakening of A antigen, and this mutation can be stably passed down to future generations.

Serological characteristics of ABO blood group and molecular genetic analysis of a Chinese pedigree with cisAB09 subtype / 中华医学遗传学杂志

OBJECTIVE@#To explore the serological characteristics of ABO blood group and molecular genetic mechanism for a Chinese pedigree with cisAB09 subtype.@*METHODS@#A pedigree undergoing ABO blood group examination at the Department of Transfusion, Zhongshan Hospital Affiliated to Xiamen University on February 2, 2022 was selected as the study subjects. Serological assay was carried out to determine the ABO blood group of the proband and his family members. Activities of A and B glycosyltransferases in the plasma of the proband and his mother were measured with an enzymatic assay. Expression of A and B antigens on the red blood cells of the proband was analyzed by flow cytometry. Peripheral blood samples of the proband and his family members were collected. Following extraction of genomic DNA, exons 1 to 7 of the ABO gene and their flanking introns were sequenced, and Sanger sequencing of exon 7 was carried out for the proband, his elder daughter and mother.@*RESULTS@#The results of serological assay suggested that the proband and his elder daughter and mother had an A2B phenotype, whilst his wife and younger daughter had an O phenotype. Measurement of plasma A and B glycosyltransferase activity suggested that the titers of B-glycosyltransferase activity were 32 and 256 for the proband and his mother, which were respectively below and above that of A1B phenotype-positive controls (128). Flow cytometry analysis showed that the expression of A antigen on the red blood cell surface of the proband has decreased, whilst the expression of B antigen was normal. Genetic sequencing confirmed that, in addition to an ABO*B.01 allele, the proband, his elder daughter and mother have harbored a c.796A>G variant in exon 7, which has resulted in substitution of the methionine at 266th position of the B-glycosyltransferase by valine and conformed to the characteristics of ABO*cisAB.09 allele. The genotypes of the proband and his elder daughter were determined as ABO*cisAB.09/ABO*O.01.01, his mother was ABO*cisAB.09/ABO*B.01, and his wife and younger daughter were ABO*O.01.01/ABO*O.01.01.@*CONCLUSION@#The c.796A>G variant of the ABO*B.01 allele has resulted in an amino acid substitution p.Met266Val, which probably underlay the cisAB09 subtype. The ABO*cisA B.09 allele encodes a special glycosyltransferase which can synthesize normal level of B antigen and low level of A antigen on the red blood cells.

Study of the molecular characteristics of a Bweak phenotype due to a novel c.398T>C variant of the ABO gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular mechanism for an individual with Bweak subtype.@*METHODS@#Serological methods were used to identify the proband's phenotype. In vitro enzyme activity test was used to determine the activity of B-glycosyltransferase (GTB) in her serum. The genotype was determined by PCR amplification and direct sequencing of exons 5 to 7 and flanking sequences of the ABO gene. T-A cloning technology was used to isolate the haploids. The primary physical and chemical properties and secondary structure of the protein were analyzed with the ProtParam and PSIPRED software. Three software, including PolyPhen-2, SIFT, and PROVEAN, was used to analyze the effect of missense variant on the protein.@*RESULTS@#Serological results showed that the proband's phenotype was Bweak subtype with anti-B antibodies presented in her serum. In vitro enzyme activity assay showed that the GTB activity of the subject was significantly reduced. Analysis of the haploid sequence revealed a c.398T>C missense variant on the B allele, which resulted in a novel B allele. The 398T>C variant has caused a p.Phe133S substitution at position 133 of the GTB protein. Based on bioinformatic analysis, the amino acid substitution had no obvious effect on the primary and secondary structure of the protein, but the thermodynamic energy of the variant protein has increased to 6.07 kcal/mol, which can severely reduce the protein stability. Meanwhile, bioinformatic analysis also predicted that the missense variant was harmful to the protein function.@*CONCLUSION@#The weak expression of the Bweak subtype may be attributed to the novel allele of ABO*B.01-398C. Bioinformatic analysis is helpful for predicting the changes in protein structure and function.

The Advances and Application of ABO Blood Group Genotyping Technology --Review / 中国实验血液学杂志

The ABO blood group system is the most important blood group system in clinical transfusion. Serological technology is a routine method for the identification of ABO blood groups, however, which have some limitations in the identification of complicated ABO samples with weakened antigens or antibodies, abnormal plasma proteins, polyagglutination, or cold agglutinin, etc. With the development of molecular biology technology, ABO blood group gene was cloned, and ABO blood group genotyping technology based on DNA was established. The genotyping technologies with different throughputs such as PCR-SSP, Droplet-AS-PCR, PCR-RFLP, PCR-SBT, SNaPshot, MALDI-TOF MS and NGS have emerged. Genotyping has overcome the limitations of serology, and has become an indispensable method to solve difficult blood type, providing strong support for the correct identification of ABO blood group, and providing guarantee for precision blood transfusion. This review summarizes the progress and application of ABO blood group genotyping methods.

Molecular analysis of 23 cases of B subgroup / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular reasons of weak expression of B antigen on the red cell.@*METHODS@#Serological test for blood group was carried out, including red cell and plasma grouping, and anti-A1 and anti-H testing, and confirming weak A or B antigens by adsorption and elution. Exons 1-7 were sequenced directly, and one of them was cloned and sequenced.@*RESULTS@#All of the 23 samples showed the weak B antigen by serological method. The alleles of the subgroups were identified by DNA sequencing, including 2 Bel subgroup, 4 B3 subgroup, 14 Bw subgroup, 2 CisAB subgroup and a novel allele. The novel allele showed a nucleotide substitution 662G>A in the exon 7, and the sequence was submitted to Blood Group Antigen Gene Mutation Database, and the novel allele was named Bel10.@*CONCLUSION@#Nucleotide substitution in exon results in blood subgroup, which showed that the antigens were weakened, and Bw phenotype was the most frequently subgroup.

Identification of a novel FUT1 allele in a Chinese individual featuring para-Bombay phenotype / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for an individual with a para-Bombay phenotype.@*METHODS@#A proband with mismatched forward and reverse serotypes for the ABO blood group was identified. Weakly expressed ABH blood type antigen on the surface of red blood cells was verified by absorption and release test, and the blood group substances in saliva was detected by sialic acid test. Exons 6 and 7 of the ABO gene and exons of the FUT1 and FUT2 genes were subjected to direct sequencing.@*RESULTS@#The proband was found to be of O type by forward ABO serotyping and AB type by reverse ABO serotyping, though H and substance A and B were detected in her saliva. DNA sequencing revealed that she has harbored c.35C/T, c.328G/A, and c.504delC compound heterozygous variants of the FUT1 gene. Haploid analysis showed that her FUT1 genotype was h328A/h35T+504delC, which has been uploaded to the NCBI website (No. MW323551).@*CONCLUSION@#The para-Bombay phenotype of the proband may be attributed to the novel compound heterozygous variants including c.504delC of the FUT1 gene, which may affect its function by altering the activity of FUT1 glycotransferase.

Serological Identification and FUT1 Gene Mutation Analysis of 8 Individuals with Para-Bombay Phenotypes in Guangxi / 中国实验血液学杂志

OBJECTIVE@#To study the serological characteristics and molecular biological basis of 8 individuals with Para-Bombay phenotypes in Guangxi area.@*METHODS@#Serological tests were used to identify the blood groups of red cells. Molecular biological methods, including PCR-SSP for ABO genotyping and DNA sequencing for FUT1, were used to detect the genotypes of ABO and FUT1 which determined the expression of H antigen.@*RESULTS@#Eight individuals in the study were all the Para-Bombay phenotypes, including 4 cases of B@*CONCLUSION@#There are varieties of molecular genetic mechanisms for Para-Bombay phenotypes. In this study, the FUT1 mutations that cause Para-Bombay phenotypes in Guangxi area are mainly h3, h

Sequence analysis for a case with Para-Bombay AB blood type / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular basis for a rare case with Para-Bombay AB blood type.@*METHODS@#Serological method was used to determine the blood type of the proband. Exons 6 and 7 of the ABO gene and the coding regions of FUT1 and FUT2 genes were analyzed by direct sequencing.@*RESULTS@#Serological results showed that the proband was a Para-Bombay AB subtype. His genotype was determined as ABO*A1.02/B.01. The proband was also found to harbor c.551-552delAG and c.881-882delTT of the FUT1 gene. For his four children, there were three type B and one type A, though the expression of the H type was normal.@*CONCLUSION@#The double deletions in the coding region of the FUT1 gene probably underlay the Para-Bombay blood type in the proband. Carrier of single-strand deletions may have a normal ABO phenotype.

Serological and molecular studies of a rare A subgroup / 中华医学遗传学杂志

OBJECTIVE@#To determine the genotype of an individual suspected for Aw through DNA sequencing.@*METHODS@#Serologic testing was carried out with standard methods. Exons 6 and 7 of the ABO genes were amplified by PCR and subjected to direct sequencing or sequenced after gene cloning.@*RESULTS@#Serological testing showed that the forward typing and reverse typing were Aw and A, respectively. DNA sequencing revealed that the individual has carried an Aw allele and an O allele. Haplotype sequencing of each allele has revealed a nt543 variant (543G>C) in the Aw allele.@*CONCLUSION@#The individual was verified as a rare A subtype, which was previously unreported in mainland China.

Identification of B Antigen Weak Expression Caused by 5873CT Mutation in Blood Group Gene Based on Sequencing Technique / 中国实验血液学杂志

OBJECTIVE@#To explore the role and significance of blood group genotyping and gene sequencing technology in the identification of blood group subtypes.@*METHODS@#Blood type of the proband and his son were identified by blood type serology, and ABO genotyping and DNA sequencing were performed according to the results of serological expression pattern.@*RESULTS@#The weak B antigen expression was found in the proband and his son by serological test, and was preliminarily identified as B3 subtype. The ABO blood group genotyping confirmed that the genotype of the proband and his son was B/O1 and B/O2, respectively. Finally, through gene sequencing, it was confirmed that the B101 allele of the proband and his son showed a heterozygous mutation of 5873CT.@*CONCLUSION@#The combination of serology, genotyping and sequencing showed find new blood group gene mutation sites, which is important strategic significance for accurate blood group identification, personalized blood use and trasfusion safety, which is beneficial to clarify the molecular biological basis of ABO blood group subtypes.

Different Subtypes Caused by c.721C>T Substitution in the Exon 7 of ABO Gene / 中国实验血液学杂志

OBJECTIVE@#To analyze the different subtypes caused by c.721C>T substitution in the exon 7 of the ABO gene, and to investigate the related molecular mechanism of different antigens expression.@*METHODS@#ABO subtypes in 7 samples were identified by standard serological methods. The exons 6, 7, and adjacent intron of ABO gene were amplified by Polymerase Chain Reaction (PCR), and the PCR products were analyzed by direct DNA sequencing and cloning sequencing.@*RESULTS@#ABO subtypes phenotypes were A@*CONCLUSION@#c.721C>T substitution in the ABO gene causes p.Arg241Trp exchange resulting in the decreasing of GTA or GTB activities and weaker antigen expression. O.01.07 is a null allele which cannot form a functional catalytic enzyme has no effect on A

Analysis of two novel variants of FUT1 gene in a Chinese family with para-Bombay phenotype / 中华医学遗传学杂志

OBJECTIVE@#To study rare para-Bombay blood type Bm@*METHODS@#ABO and H phenotype of the proband and her pedigree were determined with serological methods. The ABO genotype was analyzed by polymerase chain reaction-sequence specific primer(PCR-SSP). The full coding region of alpha-l,2 fucosyltransferase (FUT1) gene of the pedigree was analyzed by polymerase chain reaction and direct sequencing of the amplified fragments. The haplotype of the FUT1 gene were analyzed by cloning sequencing.@*RESULTS@#The rare para-Bombay blood type Bm@*CONCLUSION@#Two new alleles of FUT1 gene (h

Serological and molecular biological analysis of an individual with para-Bombay blood group due to homozygous c.948C>A variant of FUT1 gene / 中华医学遗传学杂志

OBJECTIVE@#To study the serological, molecular and genetic characteristics of an individual with para-Bombay blood group.@*METHODS@#Serological method was used to detect the presence of A, B, H antigens in red blood cells and saliva, and Sanger sequencing was used to analyze the FUT1 gene of the proband and her family members. Genetic mechanism of the blood group was analyzed by pedigree analysis.@*RESULTS@#Forward and reverse typing of the ABO blood group were inconsistent for the proband. A, B and H antigens were not found on erythrocytes, while B and H antigens were found in saliva, in addition with unexpected antibodies. The proband was found to have a genotype of ABO*B.01/ABO*O.01.04 caused by homozygous variant of c.948C>A (p.Tyr316Ter) of the FUT1 gene.@*CONCLUSION@#A novel para-Bombay blood group was identified, which was due to the missense variant of c.948C>A in the coding region of the FUT1 gene, which has probably resulted in inability to synthesis active H antigen transferase.

Identification and pedigree analysis for an A(W)37B subtype due to c.940A>G variant of ABO gene / 中华医学遗传学杂志

OBJECTIVE@#To delineate the serological and molecular profiles of a patient with A(w)37B subtype.@*METHODS@#The ABO bloodtypes of the proband, his wife and daughter were determined with a standard serological method. Their ABO genotypes were determined by sequence-specific primer polymerase chain reaction (PCR-SSP). All exons of the ABO gene were directly sequenced. Exons 6 and 7 of the ABO gene were further analyzed by cloning and sequencing.@*RESULTS@#The red blood cells of the proband showed a weak B phenotype. His serum sample contained weak reactive anti-A antibody, which was defined as A(w)B blood group based on the serological characteristics. The A and B alleles were detected by blood group genotyping. Gene cloning and sequencing have identified a characteristic c.940A>G variant (ABO*AW.37) in exon 7 of the ABO gene, which resulted in substitution of Lysine by Glutamate at position 314. The proband's daughter has inherited the ABO*AW.37 allele.@*CONCLUSION@#The c.940A>G variant in exon 7 of the ABO gene probably underlay the decreased activity of GTA transferase and resulted in the Aw37 phenotype.

Identification of a glycosyltransferase allele associated with Bw subtype and analysis of the protein structure / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular basis for an individual with Bw subtype.@*METHODS@#Routine serological reactions were used to determine the surface antigens of erythrocytes and antibodies in serum. PCR-sequence-based typing (PCR-SBT) was used to analyze the coding regions of the ABO gene and erythroid-specific regulatory element in its intron 1. Amplicons for exons 5 to 7 containing the variant site were subjected to TA cloning for the isolation of the haploid and verification of the sequence. The 3D structure of mutant protein was predicted with Pymol software. Changes of amino acid residues and structural stability were also analyzed.@*RESULTS@#Serological assay showed that the individual had weakened B antigen and anti-B antibody in his serum. His genotype was determined as ABO*B.01/ABO*O.01.01. Sequencing of the entire coding region of the ABO gene identified an additional heterozygous c.734C/T variant. No variant was found in the erythroid-specific regulatory element of intron 1. Haploid cloning and isolation has obtained an ABO*O.01.01 allele and a ABO*B.01 allele containing a c.734T variant, which has led to substitution of Thr by Ile at position 245 in the functional center of glycosyltransferase. Based on the 3D structure of the protein, the residues binding with the mutation were unchanged, but the bonding distance between the hydrogens was changed with the amino acid substitution. Meanwhile, the connections with water molecules were increased.@*CONCLUSION@#The c.734C>T variant of the GTB gene can lead to an amino acid substitution in the functional center of the enzyme, which in turn may affect the stability of glycosyltransferase B protein and reduceits enzymatic activity.

Study of a Chinese pedigree carrying a novel variant of α-1, 3-N-acetyl galactosaminyl transferase gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a Chinese pedigree with a novel ABO subtype.@*METHODS@#The proband and his family members were subjected to serological analysis, and their genotypes were determined by fluorescence PCR and direct sequencing of the coding regions of the ABO gene. Exons 6 to 7 of the ABO gene were also subjected to clone sequencing for haplotype analysis.@*RESULTS@#The proband was determined as an AxB subtype. By fluorescence PCR, he was typed as A/B. Clone sequencing has revealed a insertional mutation c.797_798 insT in exon 7 of the ABO gene, which yielded a novel allele. Pedigree analysis confirmed that the novel ABO*A1.02 allele carried by the proband and his sister was inherited from their father. The c.797_798insT mutation has been submitted to GenBank with an accession number of MK125137.@*CONCLUSION@#The c.797_798insT mutation of exon 7 of the ABO gene probably has led to weakened expression of A antigen.

Investigation of ABO allelic competition phenomena in a pedigree with Bw11 subtype / 中华医学遗传学杂志

OBJECTIVE@#To investigate the serological and molecular characteristics of a pedigree carrying an allele for ABO*BW.11 blood subgroup.@*METHODS@#The ABO blood type of 9 pedigree members were determined by serological methods. Exons 6 and 7 of the ABO gene were amplified by PCR and directly sequenced. The patient and her father were also subjected to clone sequencing analysis.@*RESULTS@#Serological tests demonstrated that the proband and her younger brother had an ABw subtype, whilst her father and two daughters had Bw subtype. Clone sequencing found that the exon 7 of the ABO gene of the proband had a T>C substitution at position 695, which was identified as a BW.11 allele compared with the reference sequence B.01. This BW.11 allele was also identified in the proband's father, brother and two daughters. Due to allelic competition, the A/BW.11 and BW.11/O alleles demonstrated significantly different phenotypes.@*CONCLUSION@#The c.695T>C substitution of the ABO gene may lead to allelic competition in the Bw11 subtype. Combined molecular and serological methods is helpful for precise blood grouping.

Serological and molecular study of a novel B(A) allele with multiple missense mutations / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular basis for an individual suspected as AwB subtype through DNA sequencing.@*METHODS@#ABO serology was carried out with the standard tube method. To identify the ABO gene haplotype, the amplicons of exon 7 were cloned and sequenced.@*RESULTS@#Serological results showed that the forward typing was AwB and the reverse typing was B. Sequencing analysis revealed that the sample has contained an O01 allele in addition with c.297A>G, c.657C>T, c.796C>A, c.803G>C, c.930G>A variants as compared with the A101 allele.@*CONCLUSION@#Through sequencing analysis, the sample with an AwB subtype by serological testing was identified as a novel B(A) phenotype, which was unreported previously.

Molecular characterization of a recombination allele of ABO blood group / 中华医学遗传学杂志

OBJECTIVE@#To analyze the molecular characteristics of a recombinant allele of the ABO blood group.@*METHODS@#The ABO phenotype was determined with the tube method. The coding regions of the ABO and FUT1 genes were analyzed by PCR-sequence based typing. The ABO alleles of the proband were determined by allele-specific primer sequencing. The full sequences of the ABO gene of the proband and her mother were determined through next generation sequencing.@*RESULTS@#The red blood cells of the proband did not agglutinate with anti-H, and the sequence of the FUT1 gene was homozygous for c.551_552delAG.The proband was thereby assigned as para-Bombay. Bi-directional sequencing also found that she was heterozygous for c.261G/del,467C>T,c.526C>G,c.657C>T,c.703G>A,c.796C>A,c.803G>C and c.930G>A of the coding regions of the ABO gene. Allele-specific primer sequencing also found her to carry a ABO*A1.02 allele and a recombinant allele from ABO*O.01.01 and ABO*B.01. The recombination site was located between nucleotide c.375-269 and c.526, and the allele was maternally derived.@*CONCLUSION@#An recombinant allele of the ABO gene has been identified, which has originated from recombination of ABO*O.01.01 with the ABO*B.01 allele.

Asociación del grupo sanguíneo ABO con susceptibilidad a COVID-19 / Association of ABO blood group with COVID-19 susceptibility

La pandemia de COVID-19, causada por el virus SARS-CoV-2, ha infectado ya a más de 25 millones de personas, ocasionando más de 850,000 muertos y causando serios problemas en hospitales y sistemas de salud en todo el mundo. Una de las mayores dificultades que presenta la infección por SARS-CoV-2 es su gran variación en presentación clínica, que puede ir desde casos asintomáticos hasta síndromes de distrés respiratorio agudo, fallo múltiple de órganos y muerte. De aquí la importancia del estudio de factores demográficos, clínicos y genéticos que permitan la identificación de personas con mayor riesgo de adquirir la infección y sufrir manifestaciones graves de la enfermedad. Un número creciente de reportes en la literatura han sugerido que el grupo sanguíneo ABO está relacionado con el riesgo a COVID-19, coincidiendo en que personas con sangre del grupo A muestran el mayor riesgo, mientras que personas con sangre del grupo O el menor. Los objetivos de esta revisión son presentar un resumen de la evidencia existente en la literatura científica reciente y discutir estas observaciones en el contexto del conocimiento sobre la asociación de los grupos sanguíneos a varias infecciones y otras enfermedades, así como de los mecanismos potenciales involucrados. Finalmente, las implicaciones de la relación entre el grupo sanguíneo y susceptibilidad a COVID-19 son también discutidas con relación a la población guatemalteca.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has already infected more than 25 million people, resulting in more than 850,000 deaths and causing serious problems in hospitals and health systems worldwide. One of the biggest problems posed by the SARS-CoV-2 infection is its great variation in clinical presentation, which can range from asymptomatic cases to syndromes of acute respiratory distress, multiple organ failure, and death. Hence the importance of studying demographic, clinical and genetic factors that allow the identification of people at increased risk of suffering serious manifestations. A growing number of reports in the literature have suggested that the ABO blood group is related to the risk of COVID-19, demonstrating that people with type A blood have the highest risk, while people with type O blood the lowest. The objective of this review is to present a summary of the existing evidence in the recent scientific literature and to discuss these observations in the context of the knowledge of the association of blood groups to various infections and other diseases, as well as the potential mechanisms involved. Finally, the implications of the relationship between the blood groups and COVID-19 susceptibility are also discussed in relationship to the Guatemalan population.