Background: Colorectal cancer is one of the most common malignancies worldwide with more than one million new cases diagnosed each year . The aim of this study is to investigate the prevalence of herpes simplex virus and Epstein-Barr virus in patients with colorectal carcinomas and polyps in comparison with healthy subjects by using the polymerase chain reaction technique.Methods: In this analytical case-control study, we selected 15 patients with colorectal cancer , 20 patients with colorectal polyps and 35 patients without malignancy as controls. Biopsy specimens were frozen under sterile conditions at -20ºC. After DNA extraction, analysis of polymerase chain reaction to detect herpes simplex virus and Epstein-Barr virus DNA in tissue samples was performed. Statistical analysis was performed with the χ2 test.Results: We observed herpes simplex DNA in 33.3% of tumor samples (5 of 15) and 20% from the non-malignant control group (7 of 35). There was no herpes simplex DNA in the polyp tissues (0 of 20). Epstein-Barr DNA was found in 60% of tumor samples (9 of 15), 35% of polyp samples (7 of 20), and 40% of the non-malignant control group (14 of 35). Statistical analysis showed no significant association between the prevalence of herpes simplex and Epstein- Barr viruses and the incidence of colorectal cancer and polyps compared with the control group.Conclusion: The results demonstrate a lack of direct molecular evidence to support an association between herpes simplex and Epstein-Barr viruses with human colorectal malignancies. These results do not exclude a possible oncogenic role of these viruses to infect different colon cells.