While the effect of fluoride on severe early childhood caries (S-ECC) is clear, knowledge of how it influences the oral microbiota and the consequential effects on oral health is limited. In this cohort study, we investigated the changes introduced in the oral ecosystem before and after using fluoride varnish in 54- to 66-month-old individuals (n=90: 18 children were sampled at 5 different time points). 16S rDNA was amplified from bacterial samples using polymerase chain reaction, and high-throughput sequencing was performed using Illumina MiSeq platforms. Many pronounced microbial changes were related to the effects of fluoride varnishing. The health-associated and were enriched in the saliva microbiome following treatment with fluoride varnishing. Co-occurrence network analysis of the dominant genera showed that different groups clearly showed different bacterial correlations. The PICRUSt algorithm was used to predict the function of the microbial communities from saliva samples. The results showed that starch and sucrose metabolism was greater after fluoride use. BugBase was used to determine phenotypes present in microbial community samples. The results showed that and (phylum ) was greater before fluoride use. We conclude that the changes in oral microbiology play a role in fluoride prevention of S-ECC.