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Nucleic acid test (NAT) blood screening for human immunodeficiency virus (HIV) and hepatitis C virus (HCV) was introduced in northern Brazil in July 2012. There are several Brazilian articles that have evaluated transfusion transmission risks for HIV and HCV. However, to our knowledge, this article is the first to evaluate the impact of HIV and HCV NAT implementation for blood screening in northern Brazil. The aim of this study was to determine the prevalence and incidence rates of HIV and HCV among blood donors and to compare the residual risk of transfusion transmission of these infections, before (2009–2011) and after (2012–2014) NAT implementation.HIV and HCV prevalence and incidence were calculated based on rates of confirmed positive samples. Residual risk estimates were based on the incidence and window model described previously. Logistic and Poisson regressions were used in the statistical analysis. A p value of not more than 0.05 was considered significant.HIV and HCV prevalence were 209.9 and 66.3 per 100,000 donations, respectively. Residual risk for HIV and HCV decreased significantly throughout the two study periods, mainly for HCV in which the reduction was one in 169,492 to one in 769,231 donations. For HIV, the decrease was one in 107,527 to one in 769,231 donations. HIV and HCV incidence rates were 21.13 and 3.06 per 100,000 persons/year before NAT and 14.03 and 2.65 per 100,000 persons/year after NAT.The HIV and HCV NAT implementation significantly increased the transfusion safety in northern Brazil, bringing benefits to recipients due to better quality of blood products produced.