Blood Group Genotyping In New Zealand: Genotyping of Kidd Blood Group System in Multiply-Transfused Patients using High Resolution Melting Analysis

Abstract

Background: Blood group genotyping is performed routinely in some countries, though this is not in New Zealand. The Kidd blood group is a simple blood group system with particular relevance to New Zealand; the rare Jk(a-b-) null phenotype is prevalent here than in many other countries. Incompatible transfusion of red blood cell products can result in haemolytic transfusion reactions. In Kidd-incompatible transfusion, this usually takes the form of delayed haemolytic transfusion reacts, occurring due to a secondary immune response. Haemagglutination techniques are in place to detect antibodies in patient’s blood and avoid incompatible transfusion. These techniques are limited and unreliable in multiply-transfused patients, the patient group most at risk of becoming immunised due to the sheer number of transfusions they receive. Purpose: To determine if introducing blood group genotyping using by high resolution melting analysis (HRMA) to detect single nucleotide polymorphisms that give rise to the various Kidd phenotypes. This was carried out through development of an in-house genotyping technique. Successful or failure of this assay would gauge whether it is feasible to introduce this method for wider blood group genotyping. Methodology: Primers were designed to flank the 838G>A and the IVS5-1g>a single nucleotide polymorphism, responsible for the Jka/Jkb antigens and the Polynesian null allele, respectively. HRMA was performed to detect the pattern of amplicon melting. This was carried out using blood samples from multiply-transfused patients. Genotyping by HRMA was confirmed by commercially available sequence specific primer (SSP) kits. This was compared to phenotype records for these patients. Results: DNA from 37 patients blood samples was successfully extracted, analysed using HRMA, confirmed using SSP and compared to phenotypes. The genotype of one patient did not match the phenotype information collected from the database. Sequencing was performed on this patient. Results supported the initial genotyping results although phenotyping could not be performed again to confirm that this was the cause of the discrepancy. HRMA results had 97% concordance with phenotype data. Conclusion: The successful implementation of HRMA genotyping indicates that it is feasible to introduce this as an in-house test to aid in the typing of multiply-transfused patients.