Introduction: Hepatitis C virus (HCV) is the leading cause of chronic liver disease, cirrhosis and hepatocellular carcinoma, as well as the most common indication for liver transplantation in many countries. Although the incidence of hepatitis C infection has decreased dramatically over the past decade, HCV infection among prisoners, especially those who are drug addicts, continues to pose a public health problem. Aim: The aim of this study is to show the prevalence of hepatitis C (or HCV infection) and risk factors in drug addict prisoners in the Penitentiary in Glina and Lepoglava. Methods: Cross-sectional research was conducted. The subjects were drug-addicted prisoners from the Lepoglava Penitentiary in Glina. The data was collected through an anonymous survey questionnaire consisting of five parts. The first part of the questionnaire refers to sociodemographic data, the second to drug use, the third to drug injection, the fourth to the experience of treatment for drug addiction and the fifth to the history of hepatitis B vaccination and exposure and testing for hepatitis C. Results: Out of a total of 160 inmates, drug addicts from two penal institutions in the Republic of Croatia, 100 respondents (response rate 62.5%), of which 52 were from the Penitentiary in Glina and 48 from the Lepoglava Penitentiary. Of the total number of participants, 25% of participants had or currently have positive hepatitis C status during their lifetime, and it is assumed that the prevalence of drug addicts with hepatitis C within the prison system covered by the survey 25%. There is an association between participation in hepatitis C education and the use of sterile needles in the last drug injection (χ2 = 4,640, p < 0.05). There is no significant association between the age of first drug use (injection) and hepatitis C status (χ2 = 0.396, p > 0.05). There is no association between hepatitis C status and the use of sterile needles in the last drug injection (χ2 = 2.048, p > 0.05). It was found that there is an association between hepatitis C status and drug use (χ2 = 5.431, p < 0.05). There was an association between involvement in a treatment program for the purpose of drug addiction and the use of sterile needles in the last injection of drugs (χ2 = 8.606, p < 0.01). Conclusion: Analysis of the obtained results showed that the prevalence of drug addicts suffering from hepatitis C within the investigated prison system is 25% and that education before staying in prison represents an important role for the use of sterile injection kits. There is no significant correlation between the age of first drug use by injection and hepatitis C, nor the habit of using sterile needles for injection in the last drug injection before being in prison. There is an association between hepatitis C status and drug use and involvement in a treatment program for the purpose of drug addiction rehab and the use of sterile utensils at the last injection before being in prison.