Alpha-thalassaemia in association with beta-thalassaemia patients in Malaysia: a study on the co-inheritance of both disorders.

BACKGROUND/AIMS: Individuals with double heterozygosity for alpha- and beta-thalassaemia and heterozygous beta-thalassaemia show a similar haematological picture. Co-inheritance of alpha- and beta-thalassaemia in both partners may result in pregnancies with either Hb Bart's hydrops foetalis or beta-thalassaemia major, or pregnancies with both disorders. METHODS: The co-inheritance of alpha-thalassaemia in 322 beta-thalassaemia carriers in Malaysia was studied. RESULTS: The frequency of alpha-thalassaemia in the beta-thalassaemia carriers was 12.7% (41/322), with a carrier frequency of 7.8% for the SEA deletion, 3.7% for the -alpha(3.7) deletion, 0.9% for Hb Constant Spring and 0.3% for the -alpha(4.2) deletion. CONCLUSION: Double heterozygosity for alpha- and beta-thalassaemia was confirmed in 5 out of the 41 couples and the risk of the fatal condition Hb Bart's hydrops foetalis was confirmed in two of these couples. Detection of the Southeast Asian (SEA) deletion in the Malaysian Malays in this study confirms that Hb Bart's hydrops foetalis can occur in this ethnic group. Results of this study have provided new information on the frequency and different types of alpha-thalassaemia (--(SEA), -alpha(3.7) and -alpha(4.2) deletions, Hb Constant Spring) in Malaysian beta-thalassaemia carriers.

Rapid and cost-effective antenatal diagnosis of haemoglobin Bart's hydrops foetalis syndrome using a duplex-polymerase chain reaction.

Haemoglobin Bart's hydrops foetalis syndrome (--SEA/--SEA) is not compatible with life and contributes to a majority of the hydropic foetuses in the Malaysian Chinese alpha-thalassaemia carriers who possess the 2-alpha-gene deletion in cis (--SEA/alphaalpha). A duplex-PCR which simultaneously amplifies a normal 136 bp sequence between the psialpha-alpha2-globin genes and a 730 bp Southeast Asian deletion-specific sequence (--SEA) between the psialpha2-theta1-globin genes was established. The duplex-PCR which detects the --SEA deletion in both chromosomes serves as a rapid and cost-effective confirmatory test in the antenatal diagnosis of Haemoglobin Bart's hydrops foetalis syndrome in Malaysia. In addition, the duplex-PCR is simple to perform as both the normal and deletion-specific alpha-globin gene sequences are amplified in the same PCR reaction.

Combine-ARMS: a rapid and cost-effective protocol for molecular characterization of beta-thalassemia in Malaysia.

Beta-thalassemia major patients have chronic anemia and are dependent on blood transfusions to sustain life. Molecular characterization and prenatal diagnosis of beta3-thalassemia is essential in Malaysia because about 4.5% of the population are heterozygous carriers for beta-thalassemia. The high percentage of compound heterozygosity (47.62%) found in beta-thalassemia major patients in the Thalassaemia Registry, University of Malaya Medical Centre (UMMC), Malaysia, also supports a need for rapid, economical, and sensitive protocols for the detection of beta-thalassemia mutations. Molecular characterization of beta-thalassemia mutations in Malaysia is currently carried out using ARMS, which detects a single beta-thalassemia mutation per PCR reaction. We developed and evaluated Combine amplification refractory mutation system (C-ARMS) techniques for efficient molecular detection of two to three beta-thalassemia mutations in a single PCR reaction. Three C-ARMS protocols were evaluated and established for molecular characterization of common beta-thalassemia mutations in the Malay and Chinese ethnic groups in Malaysia. Two C-ARMS protocols (cd 41-42/IVSII #654 and -29/cd 71-72) detected the beta-thalassemia mutations in 74.98% of the Chinese patients studied. The CARMS for cd 41-42/IVSII #654 detected beta-thalassemia mutations in 72% of the Chinese families. C-ARMS for cd 41-42/IVSI #5/cd 17 allowed detection of beta-thalassemia mutations in 36.53% of beta-thalassemia in the Malay patients. C-ARMS for cd 41-42/IVSI #5/cd 17 detected beta-thalassemia in 45.54% of the Chinese patients. We conclude that C-ARMS with the ability to detect two to three mutations in a single reaction provides more rapid and cost-effective protocols for beta-thalassemia prenatal diagnosis and molecular analysis programs in Malaysia.

High performance liquid chromatographic determination of aluminium in natural waters in the form of its lumogallion chelate.

A high performance liquid chromatographic method for the determination of ultra trace amount of aluminium in natural waters has been developed using lumogallion as a precolumn reagent for fluorimetric detection. The highly fluorescent Al-lumogallion chelate (lambda(ex) 500 nm, lambda(em) 574 nm) was separated on a LiChrosorb RP 18 column with an eluent containing 3:7 acetonitrile/0.02M potassium hydrogen phthalate buffer (pH 4.7) containing 10(-5)M lumogallion. The proposed system provides a simple, quick, selective and sensitive method for the determination of ultra-trace amount of aluminium in water samples. The detection limit defined as three times the standard deviation of the blank signal, was 0.05 mug/l. in water samples for 100 mul injection. The tolerance limits were 5 mg/l. for Fe(III) and F(-) and over 10 mg/l. for other foreign ions. The sensitivity of the method was independent of salinity. This method had been used for the direct determination of aluminium in both tap and coastal sea-waters without any preconcentration steps.

IAA-induced apogamy in Platycerium coronarium (Koenig) Desv. gametophytes cultured in vitro.

Apogamous sporophytes were produced on Platycerium coronarium gametophytes cultured in the presence of indole-3-acetic acid (IAA). The percentage of apogamy as well as the total number of apogamous sporophytes produced per gametophyte clump were highest in the presence of 40 µM IAA. When ethylene was allowed to accumulate in the culture vessel in the presence of an optimum level of IAA, the percentage and total number of apogamous sporophyte production decreased significantly. Using light microscope and confocal laser scanning microscope we have shown that nuclear size can be used as a quick parameter to estimate the ploidy level of P. coronarium.

Heavy metals in some Chinese herbal plants.

The concentrations of nine heavy metals, cadmium, cobalt, copper, iron, manganese, nickel, lead, zinc, and mercury in 42 Chinese herbal medicinal plants were determined. Generally, all the samples studied had, relative to the other trace metals, higher concentrations of iron, manganese, and zinc. The concentration range of the metals determined was comparable to that in many of the East Asian vegetables and fruits. A few samples were found to contain relatively higher concentrations of the toxic metals such as cadmium, lead, and mercury. This was probably caused by contamination during air-drying and preservation.

Production of aposporous gametophytes and calli from Pteris vittata L. pinnae strips cultured in vitro.

Murashige and Skoog's modified medium in 1% Difco Bacto-agar supplemented with sugar alcohols (sorbsitol, mannitol), growth regulators (1-naphthalenacetic acid, 2,4-dichlorophenoxyacetic acid, benzyladenine, kinetin) and sugars (fructose, glucose, sucrose) induced aposporous gametophytes from pinnae of Pteris vittata cultured in vitro at lower concentrations of all the mentioned components. Aposporous gametophytes and vegetative calli were produced at higher concentrations. The calli regenerated sporophytes when cultured on MS medium without growth regulators. The gametophytes grew vegetatively on MS medium but produced sporophytes when transferred into 0.1 strength MS medium. This is the first report of simultaneous production of calli and gametophytes from fern explants.

Production of first and second generations aposporous gametophytes fromPyrrosia piloselloides (L.) Price frond strips cultured in vitro.

Second generation aposporous gametophytes were obtained from sporophytes derived from first generation aposporous gametophytes, which in turn came from the mature fronds grown from spores in the laboratory. Murashige and Skoog modified medium in 1% agar supplemented with sugar alcohols (sorbitol, mannitol), auxins (NAA, 2,4-D) and cytokinin (BA) promoted a higher percentage of aposporous development from mature fronds ofPyrrosia piloselloides derived from aseptically cultured spores as compared with those obtained from plants in the field. A method using 4'6-diamidino-2-phenyl indole and fluorescence microscopy correlated the deoxyribonucleic acid contents of the aposporous gametophytes and sporophytes derived from them with their ploidy level.

Production of aposporous gametophytes from Drymoglossum piloselloides (L.) Presl. frond strips.

Strips of Drymoglossum piloselloides fronds in a modified Murashige and Skoog medium in 1% agar produced aposporous gametophytes in about 2 months. Young fronds showed a greater ability to develop aposporous gametophytes than older fronds. The addition of kinetin to the medium improved the ability of older fronds while the presence of 1-naphthaleneacetic acid enhanced the effect of kinetin.