Impact of in vitro inoculation and dietary supplementation with Bacillus subtilis on sperm quality of aged White Leghorn roosters

The probiotic Bacillus subtilis improves broiler performance; however, its effects on rooster reproduction are unknown. Therefore, 2 experiments (EXP) were conducted to evaluate impacts of B. subtilis on poultry semen quality. In EXP 1, B. subtilis was cultured for 48 h to a concentration of 10⁸ CFU/mL. Semen from 72-week-old White Leghorn roosters was pooled and diluted 10-fold with the following treatments: 1) saline control, 2) sterile broth, 3) culture of B. subtilis, 4) supernatant from the culture, and 5) bacterial pellet from the culture. Semen pH and the sperm quality index (SQI) were obtained at 0 and 10 min post dilution to analyze effects of exposure length. The entire experiment was replicated 3 times. Semen pH and SQI were not affected by the B. subtilis pellet as compared to saline control. However, pH and SQI for every treatment containing broth were lower than the saline or pellet treatments. For EXP 2, 42 individually caged White Leghorn roosters, 74 wk old, were fed either 0 or 4.5 × 10⁴ CFU of B. subtilis/g of feed. For each of 4 wk, individual semen samples were analyzed for pH, semen volume, sperm concentration, sperm viability, and the SQI. Additionally, semen concentrations of Na⁺, Ca²⁺, K⁺, Cl⁻, CO₂, and O₂ were measured. In the last week, ejaculates were serially diluted and plated to determine Bacillus spp. counts. The dietary addition of B. subtilis did not alter sperm quality characteristics, seminal ion concentrations, or Bacillus spp. counts. In conclusion, neither direct exposure of sperm nor dietary exposure of roosters to B. subtilis alters sperm quality, possibly because this bacterium is indigenous to the rooster's reproductive tract and semen. Therefore, feeding B. subtilis to roosters may not negatively affect fertility and may be an acceptable method to decrease pathogens, because B. subtilis alters intestinal microbiota. Future studies should investigate the effect of this probiotic on semen microbiota, especially regarding the presence of pathogenic bacteria that threaten public health.