Newborn Screening for Hemoglobinopathies and Red Cell Enzymopathies in Tripura State: A Malaria-Endemic State in Northeast India

Hemoglobinopathies are a group of inherited single gene disorders. There are reports on hemoglobin (Hb) variants identified in the tribal and non-tribal populations of Tripura State in northeastern India. This study aimed to determine the spectrum of hemoglobinopathies and enzymopathies by newborn screening in Tripura State and assess the extent of neonatal jaundice. A total of 2400?cord blood samples were collected and analyzed by high performance liquid chromatography (HPLC). Further confirmation of any abnormal HPLC was done by DNA analysis. The samples were also screened for deficiency of enzymopathies, glucose-6-phosphate dehydrogenase (G6PD) deficiency and pyruvate kinase. Of 2400?cord blood samples screened, 225 (9.3%) were Hb E (HBB: c.79G>A) heterozygotes, 80 (3.3%) were Hb E homozygotes and one carried Hb E-β-thalassemia (β-thal). Other Hb abnormalities were also detected including 15 Hb S (HBB: c.20A>T) heterozygotes, two Hb D-Punjab (HBB: c.364G>C) heterozygotes and two compound heterozygotes for Hb D-Punjab and Hb E. Of the 80 homozygous Hb E babies, four were non-tribal and 76 babies were tribal, and 225 patients carried Hb E trait, 141 were tribal, while 84 were non-tribal. Of 40 G6PD deficient babies identified, 13 had coinherited Hb E and two babies had pyruvate kinase deficiency. α Genotyping was performed in 162 affected babies, 50 of them carried α gene deletions. Newborn screening programs for Hb E, other hemoglobinopathies and G6PD deficiency must be encouraged in the malaria-endemic northeastern region of India. Drug-induced hemolysis can also be avoided by screening for G6PD deficiency at birth.     

Prevalence of the Sickle Cell Gene in Yemen: A Pilot Study

To determine the prevalence of the sickle cell gene (β^S) in Yemen and amongst people from different regions of the country living in the capital, Sana'a City, cord blood samples from 1,500 consenting mothers were collected from hospitals in Sana'a City between July and December 2001. The names and original homes of the parents were recorded. Cation exchange high performance liquid chromatography (HPLC) analysis was used for screening, while isoelectric focusing (IEF) and DNA polymerase chain reaction (PCR) were used to confirm Hb S [β6(A3)Glu→Val]. Thirty‐three samples were found to show Hb FAS, giving an overall likely β^S gene frequency of 0.011. The β^S gene frequency varied with the part of the country from which the parents came. Amongst people from Taiz and Haja in the west, the gene frequency was more than 0.04, but less than 0.004 amongst people from Ibb, adjacent to the governorate (administrative division) of Taiz. Of 66 chromosomes from babies carrying the β^S gene, only 1.5% also carried the ? 158 (C→T) ^Gγ‐globin gene XmnI site compared with 16.1% of 168 chromosomes from babies without the β^S gene from the same regions. The results of this study show a higher β^S gene frequency in the western coastal part of Yemen than in the central mountainous and eastern desert areas. The incidence of affected homozygous births may therefore reach 20/10,000 in certain areas, although it is much lower than this overall. Limited health resources can best be invested in developing a program of education, screening and health care, initially prioritizing those communities residing in the western areas of Yemen with the highest β^S gene frequency.     

Prevention and treatment of COVID-19 in patients with benign and malignant blood disorders

Patients with moderate to severe immunosuppression, a condition that is common in many hematologic diseases because of the pathology itself or its treatment, are at high risk for COVID-19 and its complications. While empirical data are sometimes conflicting, this heightened risk has been confirmed in multiple well-done studies for patients with hematologic malignancies, particularly those with B-cell lymphoid malignancies who received lymphocytotoxic therapies, those with a history of recent hematopoietic stem cell transplant and chimeric antigen receptor T-cell therapy, and, to a lesser degree, those with hemoglobinopathies. Patients with immunosuppression need to have a lower threshold for avoiding indoor public spaces where they are unable to effectively keep a safe distance from others, and wear a high-quality well-fitting mask, especially when community levels are not low. They should receive an enhanced initial vaccine regimen and additional boosting. Therapeutic options are available and immunosuppressed patients are prioritized per the NIH.     

α‐Thalassemia 2, 3.7 kb deletion and hemoglobin AC heterozygosity in pregnancy: a molecular and hematological analysis

α‐Thalassemia is a synthesis hemoglobinopathy with a worldwide distribution. α‐thalassemia‐2^3.7kb (α‐Thal2^3.7kb) was investigated by PCR and standard hematologic analysis techniques in 106 pregnant women – 53 heterozygous for hemoglobin (Hb) A and C (AC) and 53 homozygous for the normal Hb A (AA) with similar ages and race ancestry. Eleven (21%) of AC women were α‐Thal2^3.7kb heterozygous and 1 (2%) was homozygous, while 12 AA women (23%) were heterozygous. In the AA group, the MCV differed among those with normal α genes and those with α‐Thal2^3.7kb (P = 0.031). Statistical analysis of AC group patients with normal α genes and α‐Thal2^3.7kb carriers showed differences in MCV (P = 0.001); MCH (P = 0.003) and Hb C concentrations (P = 0.011). Analysis of AA and AC group patients with normal α genes showed differences in RBC (P = 0.033), Hb concentration (P = 0.003) and MCHC (P < 0.0001). There were no statistically significant differences for any hematologic parameters between AC and AA group patients with the α‐Thal2^3.7kb genotype. The AC α‐Thal2^3.7kb homozygous women had low hematologic parameters. Serum ferritin levels were normal among the groups studied. These results emphasize the importance of diagnosis and follow‐up of patients with hemoglobinopathy carriers during pregnancy in order to administer adequate therapy and avoid further complications for mothers and newborns.     

Unrelated Donor Allogeneic Hematopoietic Stem Cell Transplantation for Patients with Hemoglobinopathies Using a Reduced-Intensity Conditioning Regimen and Third-Party Mesenchymal Stromal Cells

Allogeneic hematopoietic stem cell transplantation for patients with a hemoglobinopathy can be curative but is limited by donor availability. Although positive results are frequently observed in those with an HLA-matched sibling donor, use of unrelated donors has been complicated by poor engraftment, excessive regimen-related toxicity, and graft-versus-host disease (GVHD). As a potential strategy to address these obstacles, a pilot study was designed that incorporated both a reduced-intensity conditioning and mesenchymal stromal cells (MSCs). Six patients were enrolled, including 4 with high-risk sickle cell disease (SCD) and 2 with transfusion-dependent thalassemia major. Conditioning consisted of fludarabine (150 mg/m²), melphalan (140 mg/m²), and alemtuzumab (60 mg for patients weighing > 30 kg and .9 mg/kg for patients weighing <30 kg). Two patients received HLA 7/8 allele matched bone marrow and 4 received 4-5/6 HLA matched umbilical cord blood as the source of HSCs. MSCs were of bone marrow origin and derived from a parent in 1 patient and from an unrelated third-party donor in the remaining 5 patients. GVHD prophylaxis consisted of cyclosporine A and mycophenolate mofetil. One patient had neutropenic graft failure, 2 had autologous hematopoietic recovery, and 3 had hematopoietic recovery with complete chimerism. The 2 SCD patients with autologous hematopoietic recovery are alive. The remaining 4 died either from opportunistic infection, GVHD, or intracranial hemorrhage. Although no infusion-related toxicity was seen, the cotransplantation of MSCs was not sufficient for reliable engraftment in patients with advanced hemoglobinopathy. Although poor engraftment has been observed in nearly all such trials to date in this patient population, there was no evidence to suggest that MSCs had any positive impact on engraftment. Because of the lack of improved engraftment and unacceptably high transplant-related mortality, the study was prematurely terminated. Further investigations into understanding the mechanisms of graft resistance and development of strategies to overcome this barrier are needed to move this field forward.     

In Memoriam

This paper summarizes the results on the epidemiology and molecular basis of thalassemias and other hemoglobinopathies in the Republic of Macedonia. Over the past 40 years, population surveys of more than 22,000 participants (school children and workers) from all over the country, have shown that the average incidence of β-thalassemia (thal) trait is 2.6%, ranging from less than 1% in the northeast to 10% in the south. The frequency of δβ-thal is 0.2%, while the frequency of the Swiss type of hereditary persistence of fetal hemoglobin (HPFH) is 0.3%. Screening of 9,619 newborns has shown that the frequency of α-thal trait is 1.5%, of which α-thal-2 is 1.45% and α-thal-1 is 0.05%. The molecular basis of the different forms of β-thal and other hemoglobinopathies has been completely defined. Among the Macedonians, over 450 β-thal chromosomes have been studied. Fifteen different β-thal mutations have been detected, four of which [IVS-I-110 (G→A), IVS-I-6 (T→C), IVS-I-1 (G→A), codon 39 (C→T)] account for 85% of all β-thal chromosomes. Among the Albanians, 48 β-thal chromosomes have been studied. Eight different mutations have been detected, four of which [codon 39, ?30 (T→A), IVS-I-110, IVS-I-1] account for 85% of all β-thal chromosomes. Four new mutations [?101 (C→A), ?87 (C→G), ?30, polyadenylation signal (poly A) (AATAAA→AATGAA)] have been characterized. Molecular analyses of DNA from over 20 unrelated cases with δβ-thal have shown that this condition is caused by a 13 kb deletion (Sicilian type); in two families a deletion of 18 to 23 kb (Macedonian type of δβ-thal) was discovered. Molecular analyses of α-thal in the Republic of Macedonia have shown the following types of molecular defects: 20.5 kb deletion, 17.5 kb deletion, 3.7 kb deletion, poly A mutation (AATAAA→AATGAA), and Hb Icaria [α142, Term→Lys, TAA→AAA (α2)]. The incidence of abnormal hemoglobins (Hbs) in the Republic of Macedonia is 0.4%. Three different α chain variants among 10 families, seven different β chain variants among 33 families, two γ chain variants in two newborns, one variant with an extended α chain, and Hb Lepore among 105 families, have been observed. Structural analysis of numerous cases with Hb Lepore showed that the variant was of the Washington-Boston type.     

Molecular Basis of β-Thalassemia and Other Hemoglobinopathies in Bulgaria: An Update

β-Thalassemia (thal) is relatively common in Bulgaria. Over the past 40 years population studies have been carried out in most parts of the country. Different approaches for the detection of β-thal carriers were used and a frequency from 0.5 to 19.9% was found. We have been studying β-thal in Bulgaria since 1965 and, based on our results, the average frequency is 2.5%. Here we update our results on the molecular basis of β-thal and include some unpublished data. One thousand seven hundred and fifty-two patients with signs of hemolysis were studied. Among these, 723 patients (41.3%) had β-thal or a related condition. In addition, blood samples from 875 newborn babies were studied. Eighteen different β-thal alleles were identified. The codon 39 (C→T) and IVS-I-110 (G→A) mutations occurred most frequently, and seven additional mutations were observed that were present at frequencies of 2.4 to 14.2%. This broad spectrum of β-thal alleles complicates the analysis for institutions involved in prenatal diagnosis. The frequency of α-thal is low (0.5% α-thal-1 and 1.6% for α-thal-2).     

Thalassemia Trait in Outpatient Clinics of Sana'a City,Yemen

Blood samples were collected from 699 patients attending out patient clinics in Sana'a City, Yemen, to obtain some idea of the prevalence of the thalassemias in our country. Complete blood count, hemoglobin (Hb) electrophoresis, quantitation of Hb A₂ and Hb F, and serum ferritin were determined. Microcytic, hypochromic red cells were found in 103 subjects (14.7%). Iron deficiency alone accounted for only a small proportion of these (n = 12), whereas features suggestive of β‐thalassemia (β-thal) were present in 31 patients (4.43%) and features suggestive of α-thal trait were found in 60 patients (8.6%). The study showed that thalassemia probably accounts for most red cell microcytosis in these out patient clinics, and could represent a significant health problem through births of homozygotes and compound heterozygotes with severe disorders. This pilot study should be repeated with improved technology, and extended to include globin gene analysis to define the nature of the disorders that remain poorly diagnosed.