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Clinical and epidemiologic studies on the hepatitis C virus (HCV) in the western Brazilian Amazon region are scarce. However, reports of clinical cases of hepatitis or pathologies associated to the HCV infection are frequent. In the state of Acre, there have been no studies on the population with the greatest exposure to parenteral transmission of virus infection. The objective of this study was to determine the prevalence of HCV infection among health care workers (HCWs) in this region. Of 2,338 HCWs, 646 were randomly selected for this study. The presence of antibody to HCV was determined. If these persons were antibody positive, they were tested for HCV RNA and the viral genotype was determined. The seroprevalence of antibody to HCV was 4.8% (31 of 646), and 3.7% (24 of 646) of those tested had detectable HCV RNA. Among these 24 viremic cases, HCV genotype 1 was most common (n = 16), followed by genotypes 3 (n = 6), 2 (n = 1), and an unidentified genotype. Infection with HCV (identified by a polymerase chain reaction) was more frequent among those with lower educational levels and lower incomes, those who lived for a longer period in the city of Rio Branco, those who reported intravenous use of vitamin complexes, those with a history of dental treatment, those with alcoholism, and women with history of caesarian parturition. The high prevalence of patients with HCV observed among HCWs in the city of Rio Branco and risk factors indicate the need for prevention and control programs, in addition to assistance programs, because this region is also hyperendemic for hepatitis B virus and hepatitis D virus.