Evaluation of the semen microbiome for fertility in obese men with next-generation sequencing

Objective: In the thesis study, it was aimed to examine the microbial content and diversity in semen samples of obese men, to determine the differences between infertile and fertile groups, and to investigate the effects of seminal microbiota on semen parameters, sperm DNA fragmentation (SDF), sperm chromatin condensation, and total antioxidant capacity. Material and method: Thirteen obese infertile men aged 18-55 with a BMI over 30 kg/m2 and five obese fertile men as the control group were included in the study. For microbiome analysis, the V3 and V4 regions of the 16S ribosomal RNA gene were sequenced using the amplicon sequencing method, one of the next-generation sequencing techniques. Sperm DNA fragmentation was analyzed by the TUNEL test, total antioxidant capacity was analyzed by the ELISA test and histone-rich sperm percentage was analyzed by the aniline blue staining method. Results: It was seen that the most abundant bacteria in both groups belonged to the phylum of Bacillota, Pseudomonadota, Actinomycetota and Bacteroidota. The most common bacteria at the genus level were Pseudescherichia, Staphylococcus, Paenibacillus, Streptococcus, Klebsiella, and Moraxella, which had similar distributions in both groups. Total antioxidant capacity values were 2.87 ± 0.21 in the infertile group and 2.69 ± 0.20 in the control group. Sperm DNA fragmentation was found to be higher in obese infertile men (mean: 1.39 ± 3.27) than in obese fertile men (mean: 12.10 ± 4.93). It was observed that histone-rich sperm percentage values in the control group (mean: 33.68 ± 15.65) were lower than the infertile group (mean: 48.15 ± 19.14). However, these differences were not statistically significant (p=0.127; p=0.564; p=0.172, respectively). A negative correlation was observed between the percentage of aniline-positive sperm and motility (p<0.0001), sperm concentration (p=0.0001) and total sperm count (p=0.001). It was determined that Brevibacterium, Paenibacillus, Alistipes, Lactiplantibacillus, Rhizobacter, Sphingomonas and Venlonella genera were correlated with SDF; Pantoea, Devosia, Bacteroides, Acidovorax genera were correlated with TAC, Fusobacterium genera was correlated with the percentage of aniline-positive sperm, and Corynebacterium, Hydrogenophaga, and Paenalcagenes genera were correlated with BMI. Conclusion: Bacterial species in semen may play a role in male infertility by affecting semen quality, sperm DNA fragmentation or total antioxidant capacity. Considering the relatively small size of the study sample (n=18), more meaningful results can be expected with a larger sample.