Rapid screening for glucose-6-phosphate dehydrogenase deficiency and haemoglobin polymorphisms in Africa by a simple high-throughput SSOP-ELISA method

TY - JOUR

T1 - Rapid screening for glucose-6-phosphate dehydrogenase deficiency and haemoglobin polymorphisms in Africa by a simple high-throughput SSOP-ELISA method

AU - Enevold, Anders

AU - Vestergaard, Lasse S

AU - Lusingu, John

AU - Drakeley, Chris J

AU - Lemnge, Martha M

AU - Theander, Thor G

AU - Bygbjerg, Ib C

AU - Alifrangis, Michael

N1 - Keywords: Anemia, Sickle Cell; Enzyme-Linked Immunosorbent Assay; Genetic Techniques; Genotype; Glucosephosphate Dehydrogenase; Glucosephosphate Dehydrogenase Deficiency; Hemoglobins; Humans; Mass Screening; Point Mutation; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Sensitivity and Specificity

PY - 2005

Y1 - 2005

N2 - BACKGROUND: Mutations in the haemoglobin beta-globin (HbB) and glucose-6-phosphate dehydrogenase (G6PD) genes cause widespread human genetic disorders such as sickle cell diseases and G6PD deficiency. In sub-Saharan Africa, a few predominant polymorphic variants of each gene account for a majority of these deficiencies. Examining at a larger scale the clinical importance of these independent genetic disorders, their possible association with malaria pathogenesis and innate resistance, and their relevance for antimalarial drug treatment, would be easier if an accurate screening method with limited costs was available. METHODS: A simple and rapid technique was developed to detect the most prominent single nucleotide polymorphisms (SNPs) in the HbB and G6PD genes. The method is able to detect the different haemoglobin polymorphisms A, S, C and E, as well as G6PD polymorphisms B, A and A- based on PCR-amplification followed by a hybridization step using sequence-specific oligonucleotide probes (SSOPs) specific for the SNP variants and quantified by ELISA. RESULTS: The SSOP-ELISA method was found to be specific, and compared well to the commonly used PCR-RFLP technique. Identical results were obtained in 98% (haemoglobin) and 95% (G6PD) of the tested 90 field samples from a high-transmission area in Tanzania, which were used to validate the new technique. CONCLUSION: The simplicity and accuracy of the new methodology makes it suitable for application in settings where resources are limited. It would serve as a valuable tool for research purposes by monitoring genotype frequencies in relation to disease epidemiology.

AB - BACKGROUND: Mutations in the haemoglobin beta-globin (HbB) and glucose-6-phosphate dehydrogenase (G6PD) genes cause widespread human genetic disorders such as sickle cell diseases and G6PD deficiency. In sub-Saharan Africa, a few predominant polymorphic variants of each gene account for a majority of these deficiencies. Examining at a larger scale the clinical importance of these independent genetic disorders, their possible association with malaria pathogenesis and innate resistance, and their relevance for antimalarial drug treatment, would be easier if an accurate screening method with limited costs was available. METHODS: A simple and rapid technique was developed to detect the most prominent single nucleotide polymorphisms (SNPs) in the HbB and G6PD genes. The method is able to detect the different haemoglobin polymorphisms A, S, C and E, as well as G6PD polymorphisms B, A and A- based on PCR-amplification followed by a hybridization step using sequence-specific oligonucleotide probes (SSOPs) specific for the SNP variants and quantified by ELISA. RESULTS: The SSOP-ELISA method was found to be specific, and compared well to the commonly used PCR-RFLP technique. Identical results were obtained in 98% (haemoglobin) and 95% (G6PD) of the tested 90 field samples from a high-transmission area in Tanzania, which were used to validate the new technique. CONCLUSION: The simplicity and accuracy of the new methodology makes it suitable for application in settings where resources are limited. It would serve as a valuable tool for research purposes by monitoring genotype frequencies in relation to disease epidemiology.

U2 - 10.1186/1475-2875-4-61

DO - 10.1186/1475-2875-4-61

M3 - Journal article

C2 - 16356170

SN - 1475-2875

VL - 4

SP - 61

JO - Malaria Journal

JF - Malaria Journal

ER -