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Objectives: To profile the seminal microbiome applying next generation sequencing. Methods: Semen samples of 67 men were involved in the study (21 men with and 46 men without prostatitis). Seminal microbiomes were profiled applying the method that uses combinatorial sequence tags attached to polymerase chain reaction primers that amplify the ribosomal ribonucleic acid V6 region. Amplified polymerase chain reaction products were sequenced using an Illumina paired-end protocol on HiSeq2000 platform. Results: The most abundant phylum in semen was Firmicutes, comprising nearly half of the sequences found (median 41.7%, quartiles 28.5-47.2%) followed by Bacteroidetes, Proteobacteria and Actinobacteria. The counts of lactobacilli were higher in healthy men than prostatitis patients (27% [20.2-34.6%] vs 20.2% [4.9-25.0%]; P = 0.05), especially for Lactobacillus iners. Proteobacteria comprised higher proportions in prostatitis patients than healthy men. The species richness was higher in prostatitis patients than healthy men (inverted Simpson index 13.5 AE 5.8 vs 10.3 AE 4.0). Conclusions: The semen of chronic prostatitis patients contains fewer healthsupporting lactobacilli, and has higher species diversity than that of healthy men. Firmicutes (especially lactobacilli), Bacteroidetes, Proteobacteria and Actinobacteria comprise the highest proportion of seminal microbiome.

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