Boar fecal and seminal microbiomes in relation to semen quality and probiotic supplementation in an antibiotic-free feeding program.

This study aimed to investigate the fecal and seminal microbiomes of boars in relation to semen quality and the effects of transitioning from antibiotics to probiotics within an antibiotic-free feeding regimen. A total of 21 fecal and 21 semen samples were collected from seven Duroc boars at three different time points: before antibiotic withdrawal (Day 0, serving as the control), after 48 days (Day 48), and after 96 days (Day 96). The fresh semen characteristics, bacterial culture contents, and the fecal and seminal microbiomes of boars were analyzed across the three time points. The collected semen was also diluted in an antibiotic-free semen extender and stored for 4 days. Sperm quality and bacterial culture contents were assessed daily throughout the storage period. On Day 96, sperm characteristics, including total sperm motility (-17.5 %, P = 0.024), sperm viability (-20.4 %, P = 0.002), and acrosome integrity (-25.8 %, P = 0.002), were significantly lower compared to Day 0. No significant differences were observed in total bacterial count or Lactobacillus sp. count across the three time points (P > 0.05). However, following the transition from antibiotics to probiotics, the proportion of Gram-negative bacteria in fresh semen increased from 69.5 % on Day 0-88.0 % on Day 96. In addition, the percentage of E. coli rose from 18.7 % on Day 0-76.0 % on Day 96. Over the 4-day storage period, the total bacterial count and total Lactobacillus sp. count increased (P < 0.05). The seminal microbiome analysis revealed that Enterobacterales (41.4 %) were the dominant bacterial group in boars with poor semen quality, whereas Bacteroidales (14.2 %) were predominant in those with good semen quality. A cladogram of the seminal microbiome identified Gammaproteobacteria as the bacterial biomarkers on Day 96 following the transition from antibiotics to probiotics. The alpha and beta diversity of the fecal microbiome showed significant differences between Day 0 and Day 96 (P = 0.048). However, no significant differences were observed in the alpha and beta diversity of the seminal microbiome (P > 0.05). In conclusion, the increase in Gram-negative opportunistic pathogens, including Gammaproteobacteria, Enterobacterales, and E. coli, is linked to reduced semen quality after switching from antibiotics to probiotics. However, this decline is unrelated to total bacterial or Lactobacillus sp. counts. While probiotic supplementation altered fecal microbiome diversity, it had no effect on seminal microbiome diversity or semen quality, with no connection between fecal and seminal microbiome changes.