Being widespread in all over the world, rotaviruses are known as the leading cause of acute gastroenteritis in forms of epidemic and endemic in infants and young children from 2-24 months of age. Regarding lack of vaccination for this virus, the primary aim of treatment for rotavirus gastroenteritis is replacement of fluids and electrolytes lost by vomiting and diarrhea, but these clinical procedures do not directly affect the rotavirus. The present study was conducted to investigate the antiviral direct effect of Melissa botany extract on reduction of titer and replication of rotavirus under in vitro conditions. In this study, after BSC-1 cell culture and provision of virus seed, the virus titer was determined using Reed-muench method. The extract of the plant was then provided by percolation method. The toxicity of the extracts on BSC-1 cell line and the direct effect of botany extract on viral suspension were examined at the baseline and 1 hour later. The cell was inoculated with virus. In this study, on the first step, Melissa extract in concentration of 200 micro g/ml did not have toxicity effect on BSC-1 cell line. Also direct antiviral effect of this extract on viral suspension in attachment and penetration phase of rotavirus resulted in reduction of virus titer from 10 [4.49] TCID [50] to 10 [2.66] TCID [50] . On the second step, the direct effect of botany extract within 1 hour decreased virus titer of 10 [4.49] TCID [50] to zero. The results of this study demonstrated that Melissa extract could be very effective in decreasing proliferation of rotaviruses. Therefore, this plant might be effective in treatment of rotavirus gastroenteritis