Objectives The MOSAIC study gathered data on chronic hepatitis C virus (HCV) infection and its treatment in various countries worldwide. Here we summarise patient and HCV characteristics in the Czech Republic and Slovakia. Methods MOSAIC was an observational study that included patients with chronic HCV infection untreated at the time of enrolment. Study collected and descriptively analysed patient demographics, disease stage and viral characteristics. Data were collected between February 2014 to October 2014. Results Among 220 patients enrolled, 51.4% were treatment-naive. The most prevalent HCV genotype was G1 (78.4%), followed by G3 (19.7%). Higher prevalence of G1 was found in treatment-experienced patients (94.3%) compared to treatment-naive (63.4%). Most participants (67.7%) presented viral RNA load of >= 800,000 IU/mL. Liver cirrhosis was reported in 24.5% of patients. Higher HCV RNA load and duration of HCV infection correlated with the degree of liver fibrosis. Anti-HCV interferon-based treatments were initiated in 88.2% of participants. Conclusions The study confirmed significant changes in the HCV genotypes partition with G3 genotype rapidly increasing in both countries, with possible impact on the WHO eradication initiative and treatment selection.