Delayed gut microbiome maturation and resistome expansion in infants with high-grade vesicoureteral reflux undergoing antibiotic prophylaxis

Scripts and data for reconstruction and application of the infant gut microbiome-age Random Forest model in the present study. Abstract: The development of the gut microbiome in early life is a critical determinant of long-term health, yet it can be profoundly influenced by disease and medical interventions. Here, we investigated the impact of vesicoureteral reflux (VUR) and continuous antibiotic prophylaxis (CAP) on the gut microbiome development in early infancy. Currently, CAP represents the only standard-of-care strategy for managing infants with high-grade VUR to prevent urinary tract infections. Using shotgun metagenomics, we longitudinally profiled fecal samples from 52 infants with high-grade VUR, half receiving CAP and half untreated, at baseline, 12 months, and 24 months. A microbiome age prediction model was constructed. This revealed a consistent delay in microbiome maturation in VUR infants compared to healthy references, characterized by the persistence of early colonizers and a correspondent delay in the establishment of obligate anaerobes, which are typically associated with mature gut ecosystems. This delay was more pronounced in CAP-treated infants, who also exhibited persistently reduced microbial diversity. Additionally, characterization of the gut resistome showed that CAP exposure drove a progressive enrichment of antibiotic resistance genes across multiple drug classes and promoted the expansion of multidrug-resistant strains, including members of Enterobacteriaceae and Enterococcus. Furthermore, genome-resolved metagenomics confirmed the co-localization of resistance determinants within pathogenic taxa, mostly Klebsiella and E. coli. Together, our findings indicate that VUR is associated with an intrinsically altered microbiome developmental trajectory, which is further exacerbated by CAP. These results highlight the importance of considering microbiome maturation and resistance dynamics when evaluating long-term treatment strategies in early life.