Background Malaria continues to be a major health problem in developing sub-Saharan countries including Ethiopia. Malaria is a complex disease and its local characteristics are determined by a variety of geographical, environmental, insect vector, host, and parasite factors.Methods A hospital based cross-sectional study was conducted to determine the prevalence of malaria and its possible association with hemoglobin level and ABO blood group among individuals attending Mekaneeyesus Primary Hospital, Estie District, northwestern Ethiopia. Socio-demographic variables and relevant data were collected from 390 randomly selected individuals through structured questionnaire. Then, thick and thin smears were prepared from finger pricked blood samples, stained, and examined microscopically for detection and identification of malaria parasites. ABO blood group and hemoglobin levels of the same subjects were also determined. The data generated were entered into a computer and analyzed for descriptive and logistic regression models using SPSS version 23.0. Variables with p-value < 0.05 in multivariable logistic regression were considered as explanatory variables.Results The overall prevalence of malaria was 8.5%; Plasmodium vivax (5.6%) was the most predominant, followed by P. falciparum (2.3%), and mixed infection of the two species (0.5%). In multivariate logistic regression analysis, being male (AOR = 3.48), under-five years of age (AOR = 72.84), rural residence (AOR = 2.64), and failing to use bed net (AOR =4.65) were significantly associated with the risk of malaria infection. Most (14.6%) of malaria positives cases were among individuals with blood group “A”, while the least number of cases were among subjects with blood group “O”. Individuals with blood group “A” were about four times more likelihood of getting malaria as compared to individuals with blood group “O” (AOR= 3.74). The prevalence of anemia was 23% and significantly associated with malaria (p<0.05).Conclusions Malaria in the study area is still higher than the average prevalence at country level. Therefore special attention should be given to the prevention and control strategies with the objective of increasing the awareness of local community towards malaria.