Gut microbiota immaturity with DL-endopeptidase deficiency links antibiotic use to preterm late-onset sepsis

Early antibiotic exposure increases late-onset sepsis (LOS) risk in preterm infants, potentially via gut dysbiosis. Analyzing 4,938 longitudinal fecal samples from preterm infants in China, the US, and the UK, we identified a differential pace of gut microbiota development among preterm infants. Delayed maturation correlated with over one-third of LOS risk associated with early antibiotic exposure. Deficiency of a bacterial DL-endopeptidase represented a hallmark of delayed microbiota development and correlated with elevated LOS risk. Supplementation with DL-endopeptidase-producing Enterococcus faecium or Limosilactobacillus reuteri activated the NOD2 receptor via muramyl dipeptide (MDP), regulated macrophage differentiation and polarization, restrained hyperinflammation via cylindromatosis (CYLD) induction, and protected neonatal mice from LOS. A pilot randomized controlled trial showed that L. reuteri supplementation enhanced fecal NOD2 activation in preterm infants. These findings link microbiota immaturity and reduced DL-endopeptidase activity to antibiotic exposure and LOS risk and highlight a candidate biomarker that warrants further validation for clinical translation.