• Narutchala Suwannakhon, Jittaphol Hemvuthiphan, Tanapat Pangeson, Khwanruedee Mahingsa, Arunee Pingyod, Wanwipa Bumrungpakdee, and Torpong Sanguansermsri.
• Department of Biology, School of Science, University of Phayao, Phayao, Thailand.
• Indian J Med Res. 2023 May 1; 157 (5): 447452447-452.

Background & ObjectivesNon-invasive prenatal testing (NIPT) of maternally inherited alleles of β-thalassaemia (MIB) remains to be a challenge. Furthermore, current techniques are not available for use as routine tests. NIPT for β-thalassaemia disease was developed by using a specific droplet digital polymerase chain reaction (ddPCR) assay to analyze the cell-free foetal DNA (cffDNA) derived from maternal plasma.MethodsPregnant women and their spouses who are at risk of bearing an offspring with β-thalassaemia disease from common MIB mutations (CD 41/42-TCTT, CD17A>T, IVS1-1G>T and CD26G>A) were enrolled. The ddPCR assay sets were constructed for each of the four mutations. All cell-free DNA samples were first screened for the paternally inherited β-thalassaemia (PIB) mutation. The PIB-negative samples were considered as non-disease and were not further analyzed. For PIB-positive samples, DNA fragments of 50-300 base pairs in size were isolated and purified, and further analyzed for MIB mutation. The allelic ratio between the mutant and the wild-type was used to determine the presence of MIB in cffDNA. All cases underwent a prenatal diagnosis by amniocentesis for a definite diagnosis.ResultsForty two couples at risk were enrolled. Twenty two samples were positive for PIBs. Among these 22 samples, there were 10 cases with allelic ratio >1.0 (MIB positive). All foetuses with over-represented mutant alleles were further diagnosed with β-thalassaemia disease; eight with compound heterozygous and two with homozygous mutations. The 20 PIB-negative and 12 MIB-negative foetuses were non-affected.Interpretation & ConclusionsThe results of this study suggest that NIPT utilizing the ddPCR assay can be effectively used for the screening and diagnosis of foetal β-thalassaemia in at risk pregnancies.

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