Abstract
BackgroundSickle cell disease (SCD) is an important cause of under-five mortality. Tanzania is the 5th country in the world with the highest births prevalence of SCD individuals. Significant advances in the neonatal diagnosis of SCD using rapid point-of-care testing have been made. However genetic confirmation is still required for positive cases, in uncertain cases, in multiply transfused patients, to resolve compound heterozygosity (Hb S/ β0 Thal or Hb S/ β+ thal) not uncommon in the coastal regions of East Africa and increasingly also for pre-marital counselling and potentially for future curative approaches such as gene therapy. The currently available DNA tests are prohibitively expensive. Here, we describe an easy-to-use, affordable and accurate β-globin sequencing approach that can be easily integrated within existing NBS for SCD and other haemoglobinopathies especially in Low- and Middle-income Countries.AimTo evaluate an affordable DNA technology for the diagnosis of Sickle cell disease and other haemoglobinopathies in a resource-limited setting.MethodsLaboratory-based validation study was conducted by Muhimbili University of Health and Allied Sciences and the University of Oxford involving sequencing of the entire β -haemoglobin locus using the Oxford Nanopore MinION platform. A total number of 36 Dried blood spots and whole blood samples were subjected to conventional protein-based methods (isoelectric focusing, HPLC), and/or sequenced by the Sanger method as comparators.ResultsSequencing results for SCD using the MinION were 100% concordant with those from the Sanger method. In addition, the long-read DNA sequencing method enabled the resolution of cases with unusual phenotypes which make up 1% of all children in Tanzania. The cost is £11/ sample for consumables, which is cheaper compared to other sequencing platforms.ConclusionsThis is the first report of a comprehensive single DNA assay as a definitive diagnostic test for SCD and other haemoglobinopathies. The test is fast, precise, accurate and affordable.
Highlights
Sickle cell disease (SCD) refers to a group of inherited red blood cell disorders that occur due to mutations in β-globin genes, one of which is invariably haemoglobin S (HbS), a variant produced as a result of glutamic acid to valine substitution at position 6 of the beta-globin chain, the underlying mutation being HBB: c.20A > T
The long-read Deoxyribonucleic acid (DNA) sequencing method enabled the resolution of cases with unusual phenotypes which make up 1% of all children in Tanzania
The cost is £11/ sample for consumables, which is cheaper compared to other sequencing platforms. This is the first report of a comprehensive single DNA assay as a definitive diagnostic test for SCD and other haemoglobinopathies
Summary
Sickle cell disease (SCD) refers to a group of inherited red blood cell disorders that occur due to mutations in β-globin genes, one of which is invariably haemoglobin S (HbS), a variant produced as a result of glutamic acid to valine substitution at position 6 of the beta-globin chain, the underlying mutation being HBB: c.20A > T. Worldwide it is estimated that > 300,000 babies are born annually with SCD and that these numbers will likely increase to 404,200 in 2050 [3]. Most of these babies are born in sub-Saharan Africa and India [4,5,6], where SCD contributes significantly to early childhood mortality and morbidity [7, 8]. We describe an easy-to-use, affordable and accurate β-globin sequencing approach that can be integrated within existing NBS for SCD and other haemoglobinopathies especially in Low- and Middle-income Countries
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