Purpose: The purpose of this in-vitro study was to evaluate the effect of mushroom extract and ozone gas on streptococcus mutans. Materials and methods: Suspensions of streptococcusmutans (NCTC 10449) in salt buffer were used to check the antimicrobial activity of aqueous extracts of mushroom and ozone. The test samples were divided as follow:Group I: Aqueous extract of mushroom, Group II: ozone gas, Group III: Aqueous extract of mushroom+ ozone gas. After exposure to different treatment, aliquots of the samples were spread on agar plate which was incubated at 37 C for 2 days. The number of bacterial colonies (CFU) on the plates was counted. Results: There was a significantdifference between different groups (f=31.69, p<0.001). The highest value of bacterial count was recordedfor ozone therapy (8.97±0.18), followed by mushroom extract (8.69±0.45), while the lowest value of bacterial count was recorded for mushroom extract + ozone therapy (3.36±0.12). Conclusions: mushroom extract + ozone gas show synergistic effect on streptococcus mutans.