Dissertation committee:
Τραυλού Ωραιάνθη, Ομότιμη Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Κωνσταντόπουλος Κωνσταντίνος, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Παναγιωτίδης Παναγιώτης, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Πολίτου Μαριάννα, Αναπληρώτρια Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ, Επιβλέπουσα
Γιαλεράκη Αργυρή, Αναπληρώτρια Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Βαλσάμη Σερένα, Επίκουρη Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Πιτταράς Θεόδωρος, Επίκουρος Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Summary:
ABO system along with the Rhesus system is the most diverse and immunogenic system of blood groups and the most important in transfusion therapy. Although most people are categorized in four ABO blood group the existence of subgroups complicate the transfusion. Respectively besides of RhD positive and RhD negative, there is a percentage of variant D antigen, that belong to those categories and carry the weak D, partial D and DEL phenotypes. Blood typing should take into account all expressions of the RhD antigen in order to avoid anti-D alloimmunization.
The aim of the study was to determine the frequency of ABO subgroups and variants of the Rhesus system, detect those polymorphisms in Greek population, clinical management and final characterization of these samples. For this purpose, two molecular methods, PCR-SSP and PCR-BeadChip, used to detect RHD gene variants.
Results: Of the 86 samples, 40 samples were tester for ABO molecular typing and 46 for RhD over a 3-year period. Of these, 40 samples were formulated as ΑΒΟ*A2B= 7(17,5%), subgroup AB: 3 (7,5%), ΑΒΟ*Ο/Α= 7 (17,5%), ΑΒΟ*Ο/Α2= 3 (7,5%), ΑΒΟ*Ο/ΑW(06,07,13) = 10 (25%), ΑΒΟ*Α/ΑW06= 4 (10%), ΑΒΟ*O/B= 2 (5%), ΑΒΟ*Ο/Β3=1 (2,5%), ΑΒΟ*O/O2= 2 (5%), ΑΒΟ*O/Ο45=1 (2,5%). In the second part of these study 46 samples of genomic DNA were tested for RhD, 70% of them were classified as D weak and their frequency were: RHD * weak D type 1 (n = 22), 47.8%, RHD * weak D type 3 (n = 3) 6.5%, RHD * weak D type 5 (n = 6) 13%, RHD * weak D type 11 (n = 4) 8.7%. RHD * weak D type 2 and RHD * weak D type 15 are represented once (2.2%). 5 types of D partial were detected: RHD * partial DV type 1 was detected in 3 blood donors (6.5%), whereas RHD * partial DIV, RHD * partial DVII, RHD * DAR 1.1 / type 4.2.1, RHD * type DAR 3.1 / type 4.0 were standardized once (2.2% incidence). Both molecular methods failed to characterize 2 samples, those were sequenced by the Sanger method.
In conclusion, the molecular assay of the ABO blood group system and the Rhesus CDE system is a valuable complementary method for prenatal testing and not an alternative to traditional prenatal testing methods because the molecular methods are not completely robust due to the generation of new mutations. The clinical significance of these changes is that D weak individuals belonging to types 1, 2, 3, 4.0, 4.1 and 5 can be treated as Rhesus D (+) and transfused with Rhesus D (+) red blood cells, while people with D type 4.2 to 11 and 15 should be treated as Rhesus D (-) and transfused with Rhesus D (-) red blood cells. D partial can produce a different epitope of the protein and therefore induce specific antibody production if transfected into Rhesus D negative patient. An RHD genotyping strategy, which will detect D-variants with an immunoassay and confirm D-negative blood donors with clinically significant alleles, could effectively enhance routine serological testing in Blood Donation. Finally, valuable RhD negative blood units could be saved if patients with specific RHD genotypes are transfused as RhD positive.
Keywords:
ABO, RhesusD, Molecular testing, PCR-SSP, PCR-BeadChip, Alleles frequency
