The simultaneous presence of alpha- and beta-thalassaemia alleles: a pitfall of thalassaemia screening.

OBJECTIVE: To compare the efficacy of routine haematological tests and molecular analysis in the diagnosis of double heterozygous alpha- and beta-thalassaemia. METHODS: Screening was carried out in extended family members from 125 families registered in the National Thalassaemia Registry, known to have both alpha- and beta-thalassaemia carriers. RESULTS: Eighty-three individuals from 59 families were identified to be double heterozygous for alpha- and beta-thalassaemia only upon molecular analyses. Among 40 married individuals, 1 was at 25% risk for having beta-thalassaemia major children and 6 for having Bart's hydrops pregnancies. CONCLUSION: Molecular analysis must be used for the accurate diagnosis of double heterozygous alpha- and beta-thalassaemia for proper risk ascertainment, especially in regions with a high prevalence of both types of thalassaemia.

Quantitation of hepatitis C viraemia by a competitive reverse transcription-polymerase chain reaction system.

Chronic hepatitis C infection is associated with the rapid development of cirrhosis and hepatocellular carcinoma. A quantitative assay to determine the level of hepatitis C (HCV) viraemia during treatment would be useful in determining the effect of antiviral agents. Such an assay has been developed with the principle of the method being the co-amplification of the viral genome isolated from the patient with an RNA competitor molecule (CM) using the competitive reverse transcription-polymerase chain reaction (RT-PCR). Known amounts of the CM compete for amplification with HCV RNA from the patient. To quantify each sample, 5 amplification reactions with titrated amounts of CM were performed. The CM can be distinguished from the normal HCV PCR product since it has been genetically altered to be a smaller molecule by the process of restriction digestion, ligation and reamplification. This quantitative method was used to monitor the viral load in 10 patients undergoing antiviral therapy with lymphoblastoid interferon. The level of HCV viraemia in these patients ranged from 10(9) to 10(12) genomes/ml serum. Declines in the level of viraemia were seen in 8 of the 10 patients after therapy. Since patients with low HCV viraemia levels are more likely to respond to interferon therapy in a sustained fashion, this method may also be employed to quantitate the level of viraemia in patients prior to interferon treatment, and may be an indicator of the dose and schedule of treatment. These results show that this quantitative method is useful in the monitoring of HCV viral load in patients.

Partial sequence analysis of the hepatitis C viral genome in Singapore patients.

The distribution of hepatitis C viral (HCV) genotypes in Singapore has not been previously determined. We studied the sera of 40 Singapore patients which were PCR-positive for HCV. The HCV genotypes were determined by direct sequencing of amplified sequences of the 5' non-coding region, after reverse transcription. Of the 40 samples, 35/40 (87.5%) were of HCV type 1, 2/40 (5.0%) were of type 2 and 3/40 (7.5%) were of type 3. The most common HCV genotype in this study was the type 1 genotype. Our results confirm the wide geographical distribution of HCV genotypes.