Objective To investigate the bacterial community diversity in human Demodex mites, so as to provide insights into unraveling the role of human Demodex mites in them caused infectious diseases. Methods From June to July 2023, Demodex mites were collected from the faces of college students in a university in Wuhu City using the adhesive tape method, and the V4 region of 16S ribosomal RNA (16S rRNA) gene and the internal transcribed spacer (ITS) gene of nuclear ribosomal DNA were amplified on an Illumina PE250 high-throughput sequencing platform. Sequencing data were spliced according to the overlapping relations and filtered to yield effective sequences, and operational taxonomic units (OTUs) was clustered. The diversity index of obtained OUTs was analyzed, and the structure of the bacterial community was analyzed at various taxonomic levels. Results A total of 57 483 valid sequences were obtained using 16S rRNA gene sequencing, and 159 OUTs were classified according to similarity. Then, OUTs at a 97% similarity were included for taxonomic analyses, and the bacteria in Demodex mites belonged to 14 phyla, 20 classes, 51 orders, 72 families, and 94 genera. Proteobacteria was the dominant phylum, and Vibrio, Bradyrhizobium and Variovorax were dominant genera. A total of 56 362 valid sequences were obtained using ITS gene sequencing, and 147 OTUs were obtained, which belonged to 5 phyla, 17 classes, 34 orders, 68 families, and 93 genera and were annotated to Ascomycota, Basidiomycota and Chytridiomycota, with Ascomycota as the dominant phylum, and Alternaria alternata, Epicoccum, Penicillium, and Sarocladium as dominant genera. Conclusions There is a high diversity in the composition of bacterial communities in human Demodex mites, with multiple types of microorganisms and high species abundance.