[Objective] Haikou is rich in hot spring resources and the research on the microbial diversity of hot springs is conducive to the further development and utilization of the microbial resources in Hainan hot springs. [Methods] Microbial ITS sequences and V3-V4 region of 16S rRNA gene in three hot springs (Rongyu hot spring on haidian island, S1; Happy farm hot spring in the crater, S2; and west coast Haichangliu hot spring, S3) water samples were sequenced by Illumina HiSeq high-throughput sequencing technology and were analyzed by bioinformatics. [Results] Analysis of alpha diversity shows that in the fungal community, S3 > S1 > S2, and in the bacterial community, S2 > S1 > S3. Beta diversity analysis shows that the compositions of fungal and bacterial communities in the three hot springs were significantly different; Classification analysis shows that the dominant phyla of hot spring fungal community were Ascomycota and Basidiomycota and those of hot spring bacterial community were Proteobacteria, Bacteroidetes, Thermi, Nitrospirae, Chlorobi, Firmicutes, Chloroflexi, Actinobacteria; Canonical correspondence analysis shows that temperature was the main influencing factor on fungus community in the three hot springs, and total phosphorus on bacterial community. [Conclusion] Haikou hot springs have abundant microbial resources. The microbial community composition is mainly affected by temperature for fungi and total phosphorus for bacteria.