Heinrich F. Steger

PRENATAL DIAGNOSIS OF β-THALASSEMIA MAJOR BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY ANALYSIS OF HEMOGLOBINS IN FETAL BLOOD SAMPLES

In Thailand and adjacent countries, most of the β-thalassemia genes are β0-thalassemia mutations that prevent the production of Hb A. We propose the quantitation of the Hb A fraction in fetal blood in the mid-trimester of pregnancy by automated high performance liquid chromatography as a reasonable prenatal diagnostic method to be applied in areas with limited laboratory facilities. Forty pregnant women at risk of delivering a child with β-thalassemia major were identified using an erythrocyte osmotic fragility test and quantitation of Hb A2. Cordocentesis was performed at the gestational age of 18–22 weeks and fetal blood was analyzed for hemoglobin fractions by automated high performance liquid chromatography. The β-globin gene mutations were characterized by β-globin gene sequencing. The 4 bp deletion at codons 41/42 (−TTCT) was the most frequent of the 40 β-thalassemia mutations observed (20/40 = 50%), followed by the splice site mutation IVS-I-1 (G → T) (7/40 = 17.5%), the nonsense mutation at codon 17 (A → T) (7/40 = 17.5%), the nonsense mutation at codon 35 (C → A) (3/40 = 7.5%), and the β+ -thalassemia promoter mutation at −28 (A → G) (3/40 = 7.5%). High performance liquid chromatography revealed nine fetuses which had only Hb F and no Hb A. All were homozygotes or compound heterozygotes for β0-thalassemia mutations. In the remaining 31 fetuses, a Hb A peak was present in the chromatograms. One fetus with 0.5% Hb A was a compound heterozygote for the −28 (A → G) and codons 41/42 (−TTCT) mutations. In the remaining 30 fetuses, the Hb A values ranged between 0.8 and 7.4%. Twenty of these, with a Hb A concentration of 1.82 ± 0.49% (range 0.8–2.8%), were β-thalassemia heterozygotes. The remaining 10 fetuses had Hb A values of 4.89 ± 1.47% (range 2.9–7.4%) and normal β-globin genes. The absence of Hb A in homozygotes or compound heterozygotes for β0-thalassemia mutations and the presence of measurable amounts of Hb A in heterozygotes and normal homozygotes, permits the diagnosis of fetuses expected to develop postnatal β-thalassemia major.

Screening for Alpha-Thalassemia-1 Heterozygotes in Expecting Couples by the Combination of a Simple Erythrocyte Osmotic Fragility Test and a PCR-Based Method

Objective: To develop a simple method for the prospective identification of couples at risk of homozygous α-thalassemia-1 (Hb Bart’s hydrops fetalis) in pregnancy. Methods: Antenatal care (ANC) women and their husbands were analyzed using a simple erythrocyte osmotic fragility (EOF) test and a PCR-based method for the detection of the mutation leading to α-thalassemia-1 of the Southeast Asian type (SEA). Results: Heterozygosity for the α-thalassemia-1 (SEA) deletion was found to correlate with an EOF value <60%. For a prospective screening, ANC women and their husbands are analyzed with the EOF test and only those having a value <60% were further checked by PCR. Of 2,769 cases analyzed during a 6-month period, 24 couples in which both partners are heterozygotes could be identified for genetic counseling and prenatal diagnosis. The application of the EOF test decreased the workload for PCR by ∼80%. Conclusion: Prospective screening for α-thalassemia-1 (SEA) heterozygotes in northern Thailand is becoming easier to realize by the combination of EOF test and PCR.

An investigation of the TH01 locus in a population from northern Thailand

Abstract The STR locus HUMTH01 was studied in 110 unrelated Thais from the area of Chiang Mai in North Thailand. By using PCR and vertical PAGE, six alleles were identified and the frequencies ranged from 0.005 to 0.400. The allele frequency distribution in this population showed significant differences from a Japanese population and other ethnic populations but was similiar to Asians in the USA and Australia. The genotype distribution meets Hardy-Weinberg expectations. The average power of exclusion (in no-parent and one-parent cases) and the discriminating power (DP) were calculated to be 0.3020, 0.4761 and 0.8722 respectively.

The distribution of D1S80 and VWA alleles in a Karen population from Northern Thailand.

The DIS80 and VWA loci were studied in a Karen population from Northern Thailand by the polymerase chain reaction and polyacrylamide gel electrophoresis. Twelve DIS80 and six VWA alleles were found. No deviations from the Hardy-Weinberg and linkage equilibrium were observed. The power of exclusion (PE) from the analysis of the DIS80 and VWA locus is 0.67 and 0.45, respectively, the power of discrimination (PD) is 0.95 and 0.85, respectively, with a combined PD of 0.99 and PE of 0.82.