The aim of this study was to assess the antimicrobial activity of the miswak chewing stick (Salvadora persica) in vivo, especially on streptococcus mutans and lactobacilli. The study was conducted clinically using patients' saliva and measuring the effect of miswak (chewing stick), miswak extract, toothbrush, and normal saline on mutans and lactobacilli. Forty male subjects aged 20-45 years were included in the study; there were four groups of ten subjects each. For the study, 50% of miswak extract (solution) was used. The levels of mutans streptococci and lactobacilli were measured using commercially available Vivacare line CRT (Caries Risk Test) bacteria 2 in 1 kit. The results showed there was a marked reduction of streptococcus mutans among all groups. When the groups were compared, the reduction of streptococcus mutans was significantly greater using miswak in comparison to toothbrushing (p = 0.013), and there was no significant difference for lactobacilli reduction (p = 0.147). It may be concluded miswak has an immediate antimicrobial effect. Streptococcus mutans were more susceptible to miswak antimicrobial activity than lactobacilli. Dietary intake of sugar and oral health status may be considered for controlled clinical trials with special emphasis on the antibacterial activity of miswak on cariogenic bacteria for a longer period of time. A toothbrush with and without toothpaste should be compared with miswak alone. Further research is needed with a larger sample size.