Hb L'Aquila [β106(G8)Leu→Val,CTG→GTG]: A Novel Thalassemic Hemoglobin Variant

A new β-globin variant at codon 106 (CTG→GTG), and which we named Hb L'Aquila [β106(G8)Leu→Val], was detected by DNA analysis. The proband and her father presented with the features of a mild β⁺-thalassemia (thal), confirmed by their α/β-globin chain biosynthesis ratios.     

Surface Plasmon Resonance-Based Molecular Detection of Hb S [β6(A3)Glu→Val,GAG→GTG] at the Gene Level

The surface plasmon resonance (SPR) approach, being a relatively novel biophysical method, is used to detect many different targets by biomolecular interaction. The SPR system uses optical and evanescent wave phenomenon. This approach does not need any labels, such as enzymes or isotopes, and the monitored interactions are in real time. In DNA-DNA interaction, the SPR approach is Tm-independent. Here we report our preliminary results for the molecular detection of the Hb S (GAG →GTG) mutation at codon 6 of the human β-globin gene. Our preliminary results show that the SPR approach could be applied as an inexpensive and fast routine test system for the molecular diagnosis of abnormal hemoglobins (Hbs), especially in premarital screening programs.     

Compound Heterozygosity for Hb S [β6(A3)Glu→Val,GAG→GTG] and a New Thalassemic Mutation [β132(H10)Lys→Term,AAA→TAA] Detected in a Family from West Africa

We describe a Hb S/β-thalassemia (β-thal) mutation involving an A→T transition at codon 132 of the β-globin gene. The mutation, in the heterozygous state, unlike several other mutations in exon 3, shows no signs of dominant thalassemia but those of a typical β⁰ carrier. Compound heterozygosity with Hb S [β6(A3)Glu→Val, GAG→GTG] showed a severe clinical picture.     

Hb MOLFETTA [β126(H4)Val→Leu,GTG→CTG]: A NEW,SILENT, NEUTRAL β CHAIN VARIANT FOUND IN AN ITALIAN WOMAN

We have identified a new neutral hemoglobin variant in a pregnant Italian woman, that resulted from a GTG → CTG replacement at codon 126 of the β chain, corresponding to a Val → Leu amino acid change at position β126(H4). Thermal and isopropanol stability tests were normal and there were no abnormal clinical features. Routine electrophoretic and ion exchange chromatographic methods for hemoglobin separation failed to show this variant, but reversed phase high performance liquid chromatography revealed an abnormal peak eluting near the normal β chain. No abnormal tryptic peptide was revealed on the high performance liquid chromatographic elution pattern of the total globin digest. The mutation was determined at the DNA level by amplification of the three β exons by polymerase chain reaction and direct sequencing of one exon that showed an abnormal migration on single strand conformational polymorphism analysis.     

Three New Hemoglobin Variants with Abnormal Oxygen Affinity: Hb Saratoga Springs [α40(C5)Lys→Asn (α1)], Hb Santa Clara [β97(FG4)His→Asn], and Hb Sparta [β103(G5)Phe→Val]

Abstract

β-Thalassemia (β-thal) and iron deficiency cause most microcytic anemias. Red cell indices and formulas have been established as simple, fast, and inexpensive in discrimination between these two hematological disorders in school children. However, whether these formulas could be applied to diagnose β-thal trait and iron deficiency in adult Thai subjects is unclear. The aim of this study was to examine the diagnostic accuracy of five red cell indices [red blood cell (RBC) counts, mean corpuscular volume (MCV), mean corpuscular hemoglobin (Hb) (MCH), mean corpuscular Hb concentration (MCHC), and red cell distribution width (RDW)] and nine formulas (RDW/RBC, RDW Index, Sirdah, Green and King, Mentzer, England and Fraser, Ehsani, Srivastava and Shine and Lal). Their sensitivity, specificity, positive predictive value (PPV), and negative predictive values (NPV), efficiency, and Youden’s Index were analyzed in 102 β-thal trait and 64 iron deficiency adult Thai subjects. The RDW/RBC formula proved to be the most reliable index as they had 100.0% specificity and PPV and the highest efficiency (94.58%) and Youden’s Index (91.18%), as well as high sensitivity (91.18%) and NPV (87.67%). Therefore, this formula could be used in initial discrimination of β-thal trait from iron deficiency in adult Thai subjects.     

Hb Beograd [β121(GH4)Glu→Val,GAA→GTA] in the Turkish Population

Hb Beograd [β121(GH4)Glu→Val, GAA→GTA] is a rare variant first reported in Yugoslavia and then in Turkey, Australia and New Zealand. We report two further unrelated cases from Turkey. The importance of identifying Hb Beograd at the molecular level, especially in regions where Hb D-Los Angeles [β121(GH4)Glu→Gln, GAA→CAA] is prevalent, is emphasized.     

Hb S [β6(A3)Glu→Val, GAG>GTG] and β-globin gene cluster haplotype distribution in Mauritania

Of 1050 Mauritanian blood donors screened from the two main racial groups, i.e., the Moors and Black Africans, 60 were found to carry Hb S [β6(A3)Glu→Val, GAG>GTG], giving a global frequency of 5.71%. The prevalence observed in the Black African Mauritanians (10.69%) is almost five times that found in the Moor group (2.25%). Four of the five main β(S) haplotypes were detected in this study: Senegal (77.8%), Benin (8.8%), Arab-Indian (5.5%) and Bantu (4.4%). These data showed that Hb S is a serious public health problem in Mauritania. They also confirm the ethnic heterogeneity of the Mauritanian population.     

Hb Alesha [β67(E11)Val→Met (GTG>ATG); HBB: c.202G > A] Found in a Chinese Girl

Mutations that cause destabilization of the hemoglobin (Hb) tetramer are a rare cause of hemolytic anemia. In contrast to the hemolytic anemia caused by enzyme deficiencies, a dominant mode of inheritance characterizes the unstable Hbs. Hb Alesha [β67(E11)Val→Met; HBB: c.202G>A] is caused by a G>A mutation at codon 67 of the β-globin gene, resulting in a valine to methionine substitution at helix E11. This replacement disrupts the apolar bonds between valine and the heme group, producing an unstable Hb and severe hemolysis. We report this rare hemoglobinopathy in a Chinese girl with severe hemolytic anemia, splenomegaly and frequent requirement for red blood cell (RBC) transfusions.     

Hb S [β6(A3)Glu→Val, GAG>GTG] in Mexican Mestizos: frequency and analysis of the 5' β-globin haplotype

Between 1978 and 2009, we studied 1,863 Mexican Mestizo patients with clinical data compatible with a hemoglobinopathy. Of these patients, 382 had some hemoglobin (Hb) abnormality (20.5%), 128 had a sickle cell hemoglobinopathy, representing a general frequency of 6.9%, which is similar to the percentage observed in previous studies on Mexican Mestizos. We analyzed the 5' β-globin haplotype (5'Hp) in 79 unrelated β(S) chromosomes (26 β(S)/β(S), 14 β(S)/β(Thal), nine β(S)/β(A) and four β(S)/β(D)), and four haplotypes were observed: 72.2% CAR 24.1% Benin, 2.5% Senegal and 1.2% Cameroon; the last two are reported for first time in Mexico. In some Latin American populations such as Brazil, the Bantu haplotype predominates, while in others such as Jamaica, the Benin haplotype is the most frequent, showing heterogeneity of African genes as a consequence of different regions involved in the slave trade.     

Hb S [β6(A3)Glu→Val,GAG>GTG] in Mexican Mestizos: Frequency and Analysis of the 5' β-Globin Haplotype

Between 1978 and 2009, we studied 1,863 Mexican Mestizo patients with clinical data compatible with a hemoglobinopathy. Of these patients, 382 had some hemoglobin (Hb) abnormality (20.5%), 128 had a sickle cell hemoglobinopathy, representing a general frequency of 6.9%, which is similar to the percentage observed in previous studies on Mexican Mestizos. We analyzed the 5' β-globin haplotype (5'Hp) in 79 unrelated β^S chromosomes (26 β^S/β^S, 14 β^S/β^Thal, nine β^S/β^A and four β^S/β^D), and four haplotypes were observed: 72.2% CAR 24.1% Benin, 2.5% Senegal and 1.2% Cameroon; the last two are reported for first time in Mexico. In some Latin American populations such as Brazil, the Bantu haplotype predominates, while in others such as Jamaica, the Benin haplotype is the most frequent, showing heterogeneity of African genes as a consequence of different regions involved in the slave trade.     

Hb Alperton [β135(H13)Ala→Val] Shows Decreased Oxygen Affinity

A 14-year-old male child presented with microcytosis, a known α⁺-thalassemia (α-thal) heterozygote and a hemizygous glucose-6-phosphate dehydrogenase (G6PD) deficiency. Furthermore, cation exchange high performance liquid chromatography (HPLC) revealed an additional peak eluting slightly before Hb A. The peak area of the variant was equal to that of Hb A, suggesting a β-globin variant. Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) analysis confirmed the mutation at the protein level. The variant was detectable by isoelectric focusing (IEF) or by reversed phase HPLC. DNA sequencing revealed a heterozygous mutation at codon 135 of the β gene, already described as Hb Alperton. Hb Alperton showed decreased oxygen affinity. Neither biochemical nor clinical characteristics for Hb Alperton have been reported so far.     

Characterization of a new hemoglobin variant: Hb Badalona (β31[B13]Leu→Val)

Hemoglobin (Hb) Badalona was identified in a 35-year-old Spanish female and two other family members. All affected subjects presented erythrocytosis and increased oxygen affinity (P(50): 21 mmHg). Hemoglobinopathy was not detected with electrophoretic methods. It was, however, separated and quantified by cation exchange and reverse-phase high-performance liquid chromatography. Hb Badalona accounted for 35% of the total Hb. No significant clinical symptoms were found to be related to this hemoglobinopathy. This is the first case of a Leu-->Val replacement at position beta31(B13) reported in the world literature.     

Hb Buffalo [α89(FG1)His→Gln (α1)], Observed Solely and in the Presence of an Hb S [β6(A3)Glu→Val] Heterozygosity

The hemoglobin (Hb) pattern of a 32‐year‐old Somali male living in The Netherlands, during routine diabetes mellitus monitoring, showed two more peaks in addition to the characteristic heterozygous Hb A/S pattern. A major peak of 15% faster than Hb A, and a minor one of 10.8%, overlapping Hb A₂ and the glycated Hb S_1c fraction were present. The patient was not anemic or microcytic but had a low haptoglobin level, possibly indicating a slightly elevated red blood cell (RBC) turnover. Hb S was confirmed by a sickle test and at the DNA level. The DNA sequence of the α1 gene revealed a C→G transversion at position 89, changing the local positively charged histidine to a neutral glutamine. This mutant has been previously described in a Yemenite woman and two apparently unrelated Somali males. Our case is the first showing Hb Buffalo in combination with Hb S and a G6PD deficiency, and is again observed in a Somali. No functional abnormalities associated with mutations at this amino acid residue are reported in the literature. Also, in this case no sign of any hematological abnormalities that could not be explained by the Hb S heterozygosity G6PD deficiency was found. The abnormal α chain is expressed at the expected rate and without thalassemic effect or instability. The mutated α chain seems to associate with a slight preference to the β^A (15%) rather than with the β^S counterpart. The sum of both Hb A^Buffalo and Hb S^Buffalo results in about 19–20% of total Hb. This figure is in agreement with a stable mutant of the α1 gene.     

Five Variants of the β-Globin Gene without Clinical or Hematological Effects: Hb Maryland [β47(CD6)Asp→His], Hb Kent [β37(C3)Trp→Cys], Hb Visayan [β136(H14)Gly→Cys], Hb Cutlerville [β138(H16)Ala→Val] and Hb Hornchurch [β43(CD2)Glu→Lys]

We report on five hemoglobin (Hb) β chain variants that were initially identified either by electrophoretic, chromatographic or isoelectric focusing (IEF) methods. These variants do not appear to be associated with clinical or hematological abnormalities. All variants were confirmed by DNA sequence analysis.     

The first case of Hb G-Honolulu [α30(B11)Glu→Gln (GAG>CAG); HBA2:c.91G>A] observed in association with Hb S [β6(A3)Glu→Val, GAG>GTG] in a healthy Italian child

We report the first observation of Hb G-Honolulu [α30(B11)Glu→Gln (GAG>CAG); HBA2:c.91G>A] in a Caucasian family and the first case of this variant to be found in association with Hb S [β6(A3)Glu→Val, GAG>GTG]. The proband was a healthy 4-year-old Italian boy. His chromatographic hemoglobin (Hb) pattern showed an abnormal peak having the typical retention time of Hb S (25.6% ), a second abnormal peak eluted soon after (13.6%) and a third minor peak eluted at the end of the run (6.5%). Identification of Hb variants were performed by peptide mapping using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Two abnormal peptides at m/z 765.1 and 922 were found, corresponding to the αT-4 and βT-1 peptides characteristic for Hb G-Honolulu and Hb S, respectively. The third minor abnormal peak presumably corresponded to the hybrid molecule (α(G-Honolulu)/β(S)). The concomitant presence of Hb G-Honolulu and Hb S does not seem to produce any relevant clinical manifestation.     

Two New Hemoglobin Variants: Hb Sinai-Greenspring [β34(B16)Val→Ile,GTC > ATC] and Hb Sinai-Bel Air [β53(D4)Ala→Asp,GCT > GAT]

Neonatal screening for hemoglobinopathies occasionally results in the detection of novel hemoglobin (Hb) variants. Two heterozygous infants were found with different β chain mutations, neither of which produced obvious clinical or laboratory abnormalities on routine examinations. The variants were characterized by cation exchange high performance liquid chromatography (HPLC), reversed phase HPLC, and sequencing of amplified β-globin genes. Functional studies could not be performed at this time.