Introduction: About one quarter of human immunodeficiency virus (HIV) infected persons in Serbia have also been found to be hepatitis C virus (HCV) co-infected. In the general population, HCV genotype 1 has been shown to be the most prevalent one. Here, we present the first study on the distribution of HCV genotypes among HIV/HCV co-infected patients in Serbia, in relation to epidemiological and clinical features. Material and methods: The study included HIV/HCV co-infected and a group of HCV mono-infected patients in the period 1998-2012, with collection of epidemiological, clinical, and behavioral data using a standardized questionnaire. The HCV genotyping to the level of pure genotype was performed by reverse hybridization. Results: Intravenous drug use (IDU) was found to be significantly more prevalent among the co-infected patients (p < 0.01). HCV genotype 1 was detected in 87% of patients with mono-infection, compared to 46.3% of patients with co-infection (p < 0.01); genotypes 3 and 4 were significantly more common among co-infected patients (6% and 5%, vs. 27% and 25%, respectively). Multivariate logistic regression confirmed IDU, infection with non-1 HCV genotype and HCV viral load over 5 log to be predictors of HIV co-infection. Conclusions: The HCV genotypes 3 and 4 were found to be significantly more prevalent among HIV/HCV co-infected patients in Serbia, compared to HCV mono-infected patients, but also more prevalent compared to the European HIV/HCV co-infected cohort. History of IDU represents an independent predictor of HCV genotypes 3 and 4 infection, with important implications for treatment.