Supan Fucharoen

Haplotype associated with the -31 (A-G) β⁺ -thalassemia gene in seven Thai individuals were examined and compared with that described originally in Japanese. Seven polymorphic restriction sites within β-globin gene cluster were determined using allele specific polymerase chain reaction (ASPCR) methods newly developed for rapid β⁺-globin haplotyping. A concordant result of DNA polymorphisms examined using ASPCR and conventional PCR-restriction fragment length polymorphism (PCR-RFLP) method was observed. It was found that all these seven Thai β⁺-thalassemia alleles were associated with the β-globin haplotype (+ - - - - - +), which is different from that described for a Japanese subject (- + + - + + -). This indicates two independent origins. As compared to the PCRRFLP method, β-globin haplotyping using ASPCR developed is easier, rapid, less time-consuming and requires no restriction digestion. The methods should also prove useful in population genetic study and linkage analysis of β- hemoglobinopathy.

To evaluate the results and cost-effectiveness of prenatal prevention measurement in severe thalassemia diseases at Srinagarind Hospital. Descriptive study. Antenatal care (ANC) Clinic, Srinagarind Hospital, Faculty of Medicine, Khon Kaen University. 1,498 thalassemic screened pregnant women first presenting at ANC Clinic at gestational age less than 17 weeks. Medical records of thalassemic screened pregnant women between February 2002 and February 2005 were analyzed. Those with a value of mean corpuscular volume (MCV) less than 80 fl, or positive dichlorophenol indophenol precipitation test (KKU-DCIP Clear Reagent Kit) underwent hemoglobin (Hb) typing by high performance liquid chromatography (HPLC) together with thalassemia investigation (complete blood count, MCV and Hb typing) of their husbands and to identify couples at risk of 3 severe thalassemia diseases; Hb Bart's hydrops fetalis, homozygous, -thalassemia and, -thalassemia/ Hb E disease. Then they were advised to undergo DNA analysis and, if they had fetal risk, appropriate prenatal diagnosis was offered. Number of affected fetuses detected by prenatal diagnosis. Nine hundred and ninety six pregnant women (66.49%) were positive on screening. Of these, 642 (64.46%) had thalassemia investigation done with their spouses. There were 19 couples at risk (1.27% of total screened pregnant women) for having fetal severe thalassemia disease from initial laboratory results. Most of them were, -thalassemia/ Hb E diseases. We found only 10 pregnant women (52.63%) that had undergone prenatal diagnosis. The consequent results were two affected fetuses (20%), one was Hb Bart's hydrops fetalis, and the other was, o-thalassemia/ Hb E disease. In these cases, their parents decided to discontinue the pregnancy. Our prevention program could save 1.14 million bahts for the cost of treatment in two prevented severe thalassemia cases. The prenatal prevention program of severe thalassemia disease at Srinagarind Hospital can effectively detect affected fetuses and reduce severe thalassemia disease, which is a major health problem in Thailand.

Hereditary persistence of fetal hemoglobin (HPFH) and deltabeta-thalassemia are heterogeneous disorders characterized by increased levels of Hb F in adult life. The distinction between these two conditions is not always possible from routine hematologic analyses. This study investigated the hematologic and molecular characteristics of high HbF determinants in Thailand, and describes a rapid DNA-based assay to facilitate diagnosis in a routine laboratory. A multiplex allele-specific polymerase chain reaction (PCR) system for rapid detection of three common DNA deletions causing (deltabeta)0-thalassemia and HPFH in South-east Asians was developed and used to examine the molecular basis for the high Hb F phenotypes in 273 unrelated Thai individuals. Hematologic data were recorded and correlated to the molecular findings. The multiplex PCR system was validated and results were completely concordant with those of other established methods. DNA analysis identified GgAg(deltabeta)0-thalassemia in 148 cases (54.2%), deletional HPFH-6 in 83 (30.4 %) and the deletion-inversion Ggamma(Agammadeltabeta)0-thalassemia in 22 (8.1 %) cases, while another 20 (7.3 %) subjects remained uncharacterized. Genotype-phenotype relationships are discussed. These data emphasize the high frequencies of deltabeta-thalassemia and HPFH in Thailand and the need for differential diagnostic methods since the hematologic parameters associated with the conditions are very similar and overlap. The multiplex allele-specific PCR approach should prove useful in complementing routine Hb analysis for the differential diagnosis of these three common causes of high Hb F determinants and should facilitate a program of hemoglobinopathy screening in the region.

We describe the hematological and DNA characterization of hemoglobin (Hb) E heterozygote with various forms of α-thalassemia in Thai individuals. Altogether, 202 unrelated adult subjects with Hb E heterozygotes either with or without α-thalassemia determinant were studied. The most prevalent interaction was found to be a double heterozygote for Hb E/α-thalassemia 2, followed by a double Hb E/α-thalassemia 1 and a Hb E/Hb Constant Spring (CS), even though the Hb CS was not detected. Double heterozygotes for Hb E and homozygous α-thalassemia 2 and Hb E with a compound α-thalassemia 2/Hb CS were also encountered with lower frequencies. Unexpectedly, as many as 18 cases previously diagnosed as Hb E carriers at routine Hb analysis were indeed Hb E heterozygotes with compound α-thalassemia 1/α-thalassemia 2, indicating a need for globin genotyping for accurate diagnosis. A change in Hb E level was observed which was related to a concomitant inheritance of α-thalassemia. The hematological expression of these Hb E heterozygotes with various forms of α-thalassemia, including a hitherto undescribed condition of double heterozygosity for Hb E/Hb Paksé identified in two subjects, is presented comparatively with those of the 80 cases of pure Hb E carriers. A multiplex allele-specific polymerase chain reaction (PCR) assay for simultaneous detection of Hb E and Hb CS genes is also described.

In order to provide a noninvasive prenatal diagnosis of the hemoglobin E (Hb E) related disorder, we have evaluated the possibility of identifying the fetal β^E‐globin gene in maternal plasma. The analysis was performed during 8 to 18 weeks of gestation using DNA extracted from 200 µL of plasma from pregnant women whose husbands carried Hb E. The β^E‐globin mutation in maternal plasma was detected by a nested PCR amplification followed by the Mnl I restriction analysis. The result was compared with that of routine analysis of the CVS specimens. Among the five pregnant women examined, the fetal β^E‐globin gene was identified in maternal plasma in three of them and the result was completely concordant with the conventional CVS analysis. This simple noninvasive prenatal detection of the fetal β^E‐globin gene should prove useful in a prevention and control program of Hb E/β‐thalassemia in countries where the β^E‐globin gene is prevalent. Copyright © 2003 John Wiley & Sons, Ltd.

Hemoglobin (Hb) Hekinan (α27; Glu–Asp) is a rare α‐chain variant found mainly in Japanese and Chinese whereas Hb E (β26; Glu–Lys) is common among Southeast Asians. We report a hitherto undescribed condition in which these two variants co‐segregate. The proband was a 25‐yr‐old Thai woman who was encountered with the presence of mild hypochromic microcytosis with Hb 8.2 g/dL, hematocrit (Hct) 26.0%, Mean Corpuscular Value (MCV) 68.6 fL, Mean Corpuscular Hemoglobin (MCH) 21.6 pg and Mean Corpuscular Hemoglobin Concentration (MCHC) 31.5 g/dL. Although Hb electrophoresis at alkaline pH did not show any abnormal band except Hb E in addition to Hb A, high performance liquid chromatography analysis revealed abnormal peaks at the Hb A and Hb E positions. DNA analysis of the proband revealed a GAG–GAT mutation at codon 27 of the minor α1‐globin gene for Hb Hekinan in trans to the South‐east Asian (SEA) deletional α‐thalassemia 1 determinant and a GAG–AAG mutation at codon 26 of the β‐globin gene for Hb E. She was therefore a triple heterozygote for these three anomalies. Family study identified that her mother was a double heterozygote for Hb Hekinan and Hb E without α‐thalassemia whereas her father was a classical Hb H disease patient. The genotype–phenotype relationship observed in this Thai family with complex hemoglobinopathies is presented and a simple DNA assay based on the polymerase chain reaction methodology for rapid diagnosis of Hb Hekinan is described.

The hereditary persistence of fetal hemoglobin (HPFH)‐6 is sporadically found in Thailand whereas the deletion–inversion type ^Gγ(^Aγδβ)⁰‐thalassemia is described among Indians. We report a hitherto un‐described case in which these two defects co‐segregate. He was a 3‐year‐old Thai boy who had a feature of thalassemia intermedia phenotype with the following hematologic data; Hb 8.8 g/dL, Hct 29.2%, MCV 66.9 fL, MCH 20 pg, and MCHC 30.1 g/dL. Hemoglobin analysis revealed 100% Hb F with only ^Gγ‐globin chain. Globin gene analyses demonstrated that he carried the HPFH‐6 deletion in trans to the Indian deletion–inversion ^Gγ(^Aγδβ)⁰‐thalassemia. Hematologic data of the patient was compared to those of the HPFH‐6 heterozygote found in his father, to ^Gγ(^Aγδβ)⁰‐thalassemia heterozygotes detected in his mother and sister, and to that of an unrelated Thai patient who was a compound heterozygote for the deletion–inversion ^Gγ(^Aγδβ)⁰‐thalassemia and HbE. Am. J. Hematol. 71:109–113, 2002.