Aim: The aim of this study was to investigate the prevalence of HCV genotypes in the Southern Region, Saudi Arabia. Materials & methods: A total of 76 HCV-positive individuals were selected for this study, including 34 males and 42 females, both acute and chronic patients. All HCV isolates were genotyped by direct sequencing of the 5´UTR region. The Chemistry profile, viral load and history of blood transfusion were collected from the hospital record. Results: The most common genotype was gt 4 (48.7%) followed by gt 1 (34.2%) and gt 3 (14.5%). Genotype 2 (2.6%) was only found in elderly male individuals. Patients with history of blood transfusion showed a high percentage of genotype 1 compared to the total number of all patients with genotype 1 (23% and 11% respectively).Biochemical assay showed high level of ALT particularly in genotype 4. No significant relationship was observed between HCV genotype and AST level between genotypes. The viral load was higher in HCV patients received blood transfusion than other genotypes. Conclusion: The prevalence of genotypes in this study confirmed the observation of other investigations, but no link was found between sex and genotype. There might be an association between blood donation in the past and infections with genotype 1.