Water fluoride level is unknown in many regions of Iran. Besides, only few non-controlled studies world-wide have assessed the effect of water fluoride on dental fluorosis and caries. We aimed to measure the fluoride level of 76 water supplies in 54 cities and evaluate the effect of fluoride on dental caries and fluorosis in a large multi-project study. n the first phase (cross-sectional), fluoride levels of 76 water tanks in 54 cities/villages in five provinces of Iran were randomly evaluated in five subprojects. In the second phase (retrospective cohort), 1127 middle school children (563 cohort and 564 control subjects) in the high and low ends of fluoride concentration in each subproject were visited. Their decayed, missing and filled teeth (DMFT) and fluorosis states were assessed. The data were analyzed using Chi-square, Mann-Whitney U and independent-samples t-test (α = 0.05). Mean fluoride level was 0.298 ± 0.340 mg/L in 54 cities/villages. Only eight water tanks had fluoride levels within the normal range and only one was higher than normal and the rest (67 tanks) were all at low levels.