16S rRNA amplicon sequencing of gut microbiota in adult male mice treated with long-term antibiotics during weaning

Antibiotics use in early life is increasingly scrutinized for its potential long-term effects on health, particularly its association with inflammatory bowel disease (IBD). The weaning period is a critical developmental window for the maturation of the gut microbiota, immune system, and intestinal barrier. However, the impact of early-life paternal antibiotic treatment on their offspring IBD susceptibility, and the molecular mechanisms of transgenerational effect, have not been described. A mouse model to investigate the effects of short-term (1 week) and long-term (6 weeks) treatment in the weaned period with ampicillin and cefixime, on colitis susceptibility in adulthood and the subsequent F1 generation. Both short-term and long-term antibiotic treatments during weaning significantly alleviated DSS-induced colitis in paternal male mice, with long-term treatment conferring a more pronounced protective effect. For the offspring, a protective effect against DSS-induced colitis was observed only in the F1 male mice of the long-term treated sires, but not in female offspring or the offspring of short-term treated sires. We found that this protective effect was associated with increased expression of the tight junction protein Ocln, mediated by decreased levels of miR-10b-5p and miR-200b-3p, with this trait being transmitted via sperm. Furthermore, long-term antibiotic treatment significantly altered the fecal microbiota of paternal males, increasing the abundance of certain beneficial bacteria but also promoting the enrichment of antibiotic-resistant strains. These findings provide new insights into the intergenerational effects of early antibiotic treatment and highlight the importance of the weaning period for long-term gut health and disease risk.