Human microbiota play an important role in the health of their human hosts. Recent studies have demonstrated that microbiota exist in seminal plasma. The current study aims to elucidate whether seminal microbiota exist in patients with different types of dysspermatism and whether bacterial biomarkers can be identified for them. A total of 159 study participants were recruited, including 22 patients with oligoasthenospermia, 58 patients with asthenospermia, 8 patients with azoospermia, 13 patients with oligospermia, and 58 matched healthy controls. Seminal microbiota composition was analyzed using 16S rRNA gene-based sequencing. The results showed that the composition of seminal microbiota of patients with dysspermatism differed from those of healthy controls. Comparison of the microbiota composition in semen samples from patients with different types of dysspermatism showed that microbiota in patients with asthenospermia and oligoasthenospermia were distinct from healthy controls in beta diversity (P < 0.05). Characteristic biomarkers, including Ureaplasma, Bacteroides, Anaerococcus, Finegoldia, Lactobacillus and Acinetobacter lwoffii, were identified based on LEfSe analysis. Inferred functional analysis based on seminal microbiome data further indicated the presence of potential pathogenic biomarkers in patients with asthenospermia and oligoasthenospermia. These results provided profiles of seminal microbiota exhibited in different types of dysspermatism, thus providing new insights into their pathogenesis. Human microbiota, with its diverse relationships-commensal, parasitic, mutualistic, and pathogenic-play an important role in human health. Recent studies have reported that microbiota exist in almost every part of human body-even in the endocrine niche, such as in tumors, blood, and synovial fluid 1-4. Advances in technology and new research have demonstrated that microbiota are found in seminal plasma, and play an important role in host homeostasis 5. It has been demonstrated that the presence of bacteria in sperm is associated with male infertility 6. Some bacteria in the urogenital tract may affect spermatogenesis and decrease sperm quality through various means, including decrease in sperm motility, deficiency in DNA integrity, and destruction of mitochondrial function 7. Escherichia coli, Mycoplasma genitalium, Ureaplasma urealyticum, Mycoplasma hominis, Staphylococcus aureus, and Chlamydia trachomatis are pathogens associated with male infertility 8-12. Several studies using high-throughput sequencing have demonstrated that seminal plasma has a bacterial community, which includes Lactobacillus, Pseudomonas, Prevotella, and Gardnerella, among others 13-19 .