PSI-12 Characterization of Boar Reproductive Tract Microbiome Prior to Puberty Attainment
Authors: McAnally Brooke E , Soffa Dallas R , Smith Molly S , Hickman-Brown Kyle J , Stockland Erin , Wiegert Jeffrey , Wu Guoyao , Poole Rebecca K 
Published: November 2023

Abstract
A large portion of microbiome research has focused on characterization of the gastrointestinal tract and female reproductive tract. However, minimal research has focused on establishing the bacterial community in other organs, specifically male reproductive tissues and gonads. Characterization of these microbiomes in early life could be an indicator of future semen microbiome composition and subsequent fertility status. The aim of this study was to characterize the bacterial communities of various male reproductive tissues including the testes, epididymis, seminal vesicles, prostate, bulbourethral glands, and preputial diverticulum in pre-pubertal boars. Crossbred boars (n = 4), fed a conventional diet (20% crude protein), were euthanized at approximately 2 months of age, and their intact reproductive tracts were removed. For each tissue (testes, epididymis, seminal vesicles, prostate, bulbourethral glands, and preputial diverticulum), a small incision was made before a sterile swab was rotated several times and placed into a sterile microcentrifuge tube for further analysis. A sterile swab was exposed to the environment and placed into a sterile microcentrifuge tube to act as a negative control. Samples were stored at -80°C before microbiome sequencing targeting the V4 hypervariable region of the 16S rRNA bacterial gene. Statistical analysis was conducted using the PROC GLM procedure in SAS 9.4. A greater relative abundance of the phylum Firmicutes was observed in the preputial diverticulum than the bulbourethral glands, seminal vesicles, and prostate (75.1 ± 3.6% vs. 62.3 ± 3.1%, 59.3 ± 3.1%, 58.4 ± 3.1%, respectively; P &lt; 0.05). Within Firmicutes, the relative abundance of the genus Blautia was reduced in the preputial diverticulum compared with testes, epididymis, seminal vesicles, bulbourethral glands, and prostate (0.6 ± 1.5% vs. 8.1 ± 1.3%, 8.1 ± 1.3%, 8.1 ± 1.3%, 7.6 ± 1.3%, 7.4 ± 1.3%, respectively; P &lt; 0.01). In contrast, the genus Peptoniphilus, under the phylum Firmicutes, was more abundant in the preputial diverticulum than the testes, epididymis, seminal vesicles, bulbourethral glands, and prostate (8.8 ± 1.5% vs. 0.3 ± 1.3%, 0.1 ± 1.3%, 0.2 ± 1.3%, 0.1 ± 1.3%, 0.2 ± 1.3%, respectively; P &lt; 0.01). The relative abundance of the phylum Bacteroidetes within the testes had a decreased relative abundance in comparison to the bulbourethral glands (14.2% vs. 23.6 ± 2.5%; P = 0.01), prostate (14.2% vs. 27.0 ± 2.5%; P = 0.01), and seminal vesicles (14.2% vs. 30.1 ± 2.5%; P = 0.01). The seminal vesicles (6.5% vs. 2.9 ± 0.8%; P &lt; 0.01) and prostate (5.4% vs. 2.9 ± 0.8%; P &lt; 0.05) had a greater relative abundance of the genus Bacteroides, within the phylum Bacteroidetes, than the testes. A greater relative abundance of the phylum Proteobacteria was observed in the preputial diverticulum than the bulbourethral gland, prostate, seminal vesicles, testes, and epididymis (13.5 ± 1.7% vs. 6.3 ± 1.5%, 6.5 ± 1.5%, 5.8 ± 1.5%, 5.6 ± 1.5%, 5.5 ± 1.5%, respectively; P &lt; 0.01). Based on these results, it appears the preputial diverticulum and the testes have distinct bacterial communities that differ from the male accessory sex glands including the prostate, bulbourethral glands, and seminal vesicles. Further research is needed to characterize the boar reproductive tract microbiome and determine its relationship with fertility.