Introduction: Glucose-6-phosphate dehydrogenase is a cytosolic enzyme that has a vital role inrnthe functioning and integrity of red blood cells. Lower activity of this enzyme leads to the rnoccurrence of acute hemolytic anemia after exposure to oxidative stressors like primaquine. rnAlthough primaquine is important for the radical cure of Plasmodium vivax and blocking the rntransmission of Plasmodium falciparum, thereby enhancing malaria elimination, there is a need rnto distinguish glucose-6-phosphate dehydrogenase deficient individuals and/or administer the rndrug with special care due to its hemolytic side effects. rnObjective: To determine the prevalence of Glucose-6-Phosphate dehydrogenase deficiency rnamong individuals suspected of malaria during their outpatient visit to Metehara Health Center, rnEastern Ethiopia.rnMaterials and Methods: A cross-sectional study was conducted from 01 September 2020 to 30 rnSeptember 2021. A structured questionnaire was used to collect the socio-demographic and rnclinical information of the study participants. Capillary and venous blood samples were collected rnbased on standard procedures for onsite screening tests, DBS preparation, and malaria rnmicroscopy. Data was recorded and analyzed by using SPSS version 23 software.rnResult: A total of 498 patients participated in the study; 62% (309) of them were males.rnAccording to the results of the biosensor screening test, the overall prevalence of glucose-6-rnphosphate dehydrogenase deficiency was 3.6% (18/498). There was a significant association rnbetween sex and history of previous malaria infection with glucose-6-phosphate dehydrogenaserndeficiency, P-value 0.032 and 0.02 respectively. Eleven out of the 17 (64.7%) Glucose-6-rnphosphate dehydrogenase deficient samples by bio-sensor screening test had confirmed rnmutations using sequencing. The G267+119C/T, A376T, and ChrX: 154535443 mutation types rnwere detected. rnConclusion and Recommendation: Phenotypically, the prevalence of glucose-6-phosphate rndehydrogenase deficiency was low. G267+119C/T is the predominant glucose-6-phosphate rndehydrogenase variant. In Metehara, Malaria patient management using the drug primaquinernunder close follow up for developments of any adverse effects is recommended.