A longitudinal characterization of the seminal microbiota and antibiotic resistance in yearling beef bulls subjected to different rate of gains using 16S rRNA gene sequencing and culturing

Background: Increasing evidence supports the existence of a microbial community in bovine semen, and that this seminal microbiota may influence not only the male fertility but also female reproduction. In this study, we evaluated the seminal and fecal microbiota in yearling beef bulls fed a common diet to achieve moderate (1.13 kg/d) or high (1.80 kg/d) rates of weight gain. Semen samples were collected on days 0 and 112 of dietary intervention (n = 19/group) as well as post-breeding (n = 6/group) using electroejaculation and the microbiota was assessed using 16S rRNA gene sequencing, qPCR, and culturing. The fecal microbiota was also evaluated and its similarity with seminal microbiota assessed. A subset of seminal bacterial isolates (n = 33) was screened for resistance against 28 antibiotics. Results: : A complex and dynamic microbiota was detected in bovine semen, and the community structure was affected by sampling time ( R 2 = 0.16, P < 0.001). Microbial richness increased from d 0 (ASVs, 253 ± 12) to d 112 (293 ± 14) ( P < 0.05). Microbial diversity did not change during pre-breeding but increased after breeding ( P < 0.05). Fusobacteriota, Bacteroidota, Firmicutes and Actinobacteriota were dominant in the seminal microbiota, and their abundance changed over time and following breeding ( P < 0.05). Seminal microbiota remained unaffected by the differential rate of gains, and its overall composition was distinct from fecal microbiota, with only 6% of the taxa shared between them. The fecal microbiota did not differ between the two dietary treatments at d 112 and post-breeding but changed over time from d 112 to post-breeding ( P < 0.05). A total of 364 isolates from 49 different genera were recovered under aerobic and anaerobic culturing. Among these seminal isolates were pathogenic species associated with respiratory disease, liver abscesses and reproductive infections, as well as those resistant to several antibiotics. Conclusions: : Our results suggest that bovine semen harbors a rich and complex microbiota which changes over time and in response to breeding activity but appears to be resilient to differential gains achieved via a common diet. Seminal microbiota is distinct from the fecal microbiota and harbors potentially pathogenic and antibiotic-resistant bacterial species.