Objectives: The aim of this study was to investigate the genotoxicity of waterpipe smoking in the lymphocytes of waterpipe smokers using chromosomal aberrations (CAs) assay. Materials and Methods: Fifty waterpipe smokers and 18 healthy nonsmokers volunteered to participate in the study. Additionally, 18 heavy cigarette smokers were recruited for comparison. Chromosomal aberrations (CAs) assay was used to evaluate DNA damage in the lymphocytes. Results: The results showed that similarly to cigarette smoking, waterpipe smoking significantly increased the frequencies of CAs (p < 0.01). In addition, the frequencies of CAs increased with more waterpipe use. Conclusions: Waterpipe smoking causes DNA damage to lymphocytes and the damage increases with more waterpipe use.