Intron 22 inversion real-time polymerase chain reaction detection in haemophilia A families from central South Africa

Abstract

Background. Intron 22 inversion (inv22) may account for 45% of all cases of severe haemophilia A. Haemophilia A is underdiagnosed in South Africa (SA), and owing to limited resources the genotypes of most haemophilia A patients are unknown. Objectives. To screen the haemophilia A population in central SA for inv22 using two novel detection methods. Methods. We recruited 62 participants from 27 families affected by haemophilia A in Free State and Northern Cape provinces. We screened for inv22 with our previously reported conventional polymerase chain reaction (PCR) method, as well as with a newly developed real-time PCR method. Sanger sequencing was performed to confirm the PCR results. Results. With the real-time PCR method, 10 of the severe haemophilia A patients and 3 carriers tested inv22-positive. The conventional PCR method and real-time PCR results were comparable in all but one case, where the discrepancy was attributed to sample-specific degradation. Inv22 was found in 29.4% of the severe haemophilia A population and 22.2% of the potential carriers. The inv22 status of most SA haemophilia A patients is currently unknown. The 29.4% of haemophilia A patients who were positive for inv22 was lower than the expected 45%, which could indicate a more prominent mutation than inv22 in the SA population. Conclusions. The above finding needs to be confirmed by performing comprehensive factor VIII gene ( F8 ) genotyping on the remainder of the haemophilia A patients in SA. The study contributes to genetic research in haemophilia A and lays a foundation for future research in haemophilia A genetics in SA.