|Abstract||Background: EBV is a lymphotropic herpesvirus and the causative agent of infectious mononucleosis. EBV is highly prevalent and has been linked to several malignancies. The virus is generally transferred by oral secretions and it persists as a latent infection in human B-cells. However, it can be also transmitted through blood transfusions and organ transplantations. The goal of this study was to (i) estimate the rate of infection of EBV in Qatar in healthy individuals, using serological testing, and viral load quantification, to (ii) study the correlation of EBV with demographic markers, and to (iii) study the molecular similarity of EBV isolates, using phylogenetic genotyping. Materials and methods: For estimating EBV seroprevalence, qualitative ELISA kits for detecting the following EBV antibodies were used: EBV viral capsid antigen (VCA) IgG & IgM, EBV nuclear antigen (EBNA) IgG & IgM, and early antigen (EA-D) IgG. To study the EBV viremia rate, DNA extracted from the buffy coat was subjected for viral detection and quantification using RT-PCR. For genotyping, nested PCR targeting the EBNA2 gene was used. And for further sub genotyping, the highly variable LMP-1 gene was amplified, cloned, sequenced and used for phylogenetic analysis. Results: Out of 673 analyzed samples (223 Qataris, 450 non-Qatari residents), 659 (97.9%) were EBV seropositive with different infection stages. Interestingly, 14 (2.1%) tested negative for all anti-EBV antibodies indicating no prior exposure to EBV. EBV DNA was detected in 354/673 (52.6%). Both EBV seroprevalence and viremia rate increased significantly with age. Genotyping for 51 randomly selected positive DNA samples showed that Genotype 1 was predominating, found in 36/51 (72.5%), while genotype 2 was found in 12 (23.5%) samples. Mixed infection was found in 2 (3.9%) samples. Surprisingly, Sub genotyping for 30 samples revealed that all tested clones (n=119) have the South Eastern Asia 1 (SEA1) strain. 30-bp and 69 bp deletions in the LMP-1 were also found in 10% and 13.3% samples, respectively. Conclusion: This is the first study investigating the seroprevalence, the viremia rate, and the molecular epidemiology of EBV among blood donors in Qatar. Hence, it should provide the epidemiologists, blood banks’ personnel, researchers and clinicians with EBV prevalence estimates and molecular epidemiology of EBV as a highly prevalent transfusion transmissible oncovirus.